Team:Alberta-North-RBI E/genetics

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=='''Genetics'''==
=='''Genetics'''==
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We plan to use a single metabolic pathway for the conversion of glucose to aromatics. We are currently engineering a Pseudomonas putida strain containing a “switch” system enabling the manufacture of either shikimic acid (SA) or cinnamic acid (CA) as a major product. Future embodiments of this strain will contain additional directing mechanisms whereby a multitude of derivative chemicals can be favored.  
+
We plan to use a single metabolic pathway for the conversion of glucose to aromatic chemicals. We are currently engineering a ''Pseudomonas putida'' strain containing a “switch” system enabling the manufacture of either shikimic acid (SA) or cinnamic acid (CA) as a major product. Future embodiments of this strain will contain additional directing mechanisms whereby a multitude of derivative chemicals can be favored.  
-
 
+
-
The enzymatic activities leading to the production of SA will be enhanced by the addition of extra-chromosomal copies of endogenous genes modified such that expression is constitutive. Where necessary, these genes will be further modified through site-directed mutagenesis to abolish feedback regulation. Optimization of metabolism to SA is based on the invention(s) described in US patents 5168056 and 6613552.
 
-
 
 
-
 
-
Enzymatic activities which lead from SA to phenylalanine (Phe) will be abolished by removing the underlying endogenous genes. These endogenous genes, in addition to a codon-optimized Rhodosporidium toruloides gene encoding for a phenylalanine ammonia-lysase (PAL) enzyme, will be re-introduced at an extra-chromosomal locus as modified genes containing inducible promoter/repressor “switches” allowing us to direct metabolism toward CA at will. The “switch” mechanism will require “cis” DNA elements immediately upstream of each enzyme’s coding sequence (CDS). In addition, each regulatory system (ie. induction, repression) will require a constitutively-regulated “trans” element encoded at a different locus which produces the protein component.
 
-
To enhance the yield of our desired products, two additional categories of changes will be made to our host strain. Enzymatic activities which direct metabolites away from the desired pathway will be abolished by removing the underlying endogenous genes and transporter proteins responsible for re-uptake of SA or CA from the medium will be removed in the same manner.
 
<html>
<html>
-
 
-
BioBricks to be made:
 
-
<TABLE WIDTH=632 BORDER=1 BORDERCOLOR="#00000a" CELLPADDING=7 CELLSPACING=0 RULES=GROUPS>
 
-
<COL WIDTH=197>
 
-
<COL WIDTH=65>
 
-
<COL WIDTH=58>
 
-
<COL WIDTH=254>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=6 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Part
 
-
name</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Gene/Locus</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Source</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Note</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutive
 
-
promoter</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Inducible
 
-
promoter (cis)</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Inducible
 
-
promoter (trans)</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Inducible
 
-
repressor (cis)</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Inducible
 
-
repressor (trans)</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Ribosome
 
-
binding site</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Transcription
 
-
terminator</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>n/a</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Synthetic</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>General
 
-
part</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Transketolase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>tktA</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutively
 
-
upregulated</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>DAHP
 
-
synthase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroF^FBR</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutively
 
-
upregulated, Thr326Pro = feedback insensitive</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>DHQ
 
-
synthase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroB</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutively
 
-
upregulated</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>DHQ
 
-
dehydratase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroD</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutively
 
-
upregulated</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Shikimate
 
-
dehydrogenase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroE</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Constitutively
 
-
upregulated</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Shikimate
 
-
kinase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroK</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>3-phosphoshikimate
 
-
1-carboxyvinyltransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1411</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Chorismate
 
-
synthase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1471</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Chorismatemutase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1410</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Aromatic
 
-
amino acid aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1561</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Aromatic
 
-
amino acid aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_3083</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Histidinol-phosphate
 
-
aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_0995</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=197 HEIGHT=6 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Phenylalanine
 
-
ammonia-lysase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=65 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000">&nbsp;</FONT></P>
 
-
</TD>
 
-
<TD WIDTH=58 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>R.
 
-
toruloides</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=254 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Introduced
 
-
as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
</TABLE>
 
-
<P STYLE="margin-bottom: 0.35cm"><BR><BR>
 
-
</P>
 
-
<P STYLE="margin-bottom: 0.35cm">Genes to be removed:</P>
 
-
<TABLE WIDTH=632 BORDER=1 BORDERCOLOR="#00000a" CELLPADDING=7 CELLSPACING=0 RULES=GROUPS>
 
-
<COL WIDTH=201>
 
-
<COL WIDTH=54>
 
-
<COL WIDTH=60>
 
-
<COL WIDTH=260>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=6 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Shikimate
 
-
kinase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>aroK</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>3-phosphoshikimate
 
-
1-carboxyvinyltransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1411</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Chorismate
 
-
synthase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1471</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Chorismatemutase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1410</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Aromatic
 
-
amino acid aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1561</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Aromatic
 
-
amino acid aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_3083</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Histidinol-phosphate
 
-
aminotransferase</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_0995</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed
 
-
and reintroduced as inducible</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
<TBODY>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Pyrroloquinolinequinone</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_3064</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Anthranilate
 
-
synthase component I</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_0413</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Anthranilate
 
-
synthase component II</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_0416</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Para-aminobenzoate
 
-
synthase subunit I</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>PPS_1911</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=7 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Shikimate
 
-
transporter</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>shiA</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2><I>P.
 
-
putida</I></FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
<TR VALIGN=BOTTOM>
 
-
<TD WIDTH=201 HEIGHT=6 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Cinnamic
 
-
acid transporter</FONT></FONT></FONT></P>
 
-
</TD>
 
-
<TD WIDTH=54 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000">&nbsp;</FONT></P>
 
-
</TD>
 
-
<TD WIDTH=60 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000">&nbsp;</FONT></P>
 
-
</TD>
 
-
<TD WIDTH=260 BGCOLOR="#ffffff">
 
-
<P><FONT COLOR="#000000"><FONT FACE="Calibri, serif"><FONT SIZE=2>Removed</FONT></FONT></FONT></P>
 
-
</TD>
 
-
</TR>
 
-
</TBODY>
 
-
</TABLE>
 
-
<P STYLE="margin-bottom: 0.35cm"><BR><BR>
 
-
 
-
 
-
 
<center>
<center>
-
<img src="https://static.igem.org/mediawiki/2012e/7/7a/UAlberta_E_Basic_biobricks.jpg" width="596px" height="353px"; >
+
<img src="https://static.igem.org/mediawiki/2012e/a/ae/Biobricks_together.png" ; >
</center>
</center>
</html>
</html>
-
 
 
-
 
 
-
<html>
 
-
<center>
 
-
<img src=" https://static.igem.org/mediawiki/2012e/9/9f/UAlberta_E_Constructs.jpg" width="767px" height="192px"; >
 
-
</center>
 
-
</html>
 
 +
Our metabolic engineering strategy is based on the invention of Dr. John Frost (US Patent No. 5168056, 6613552). The enzymatic activities leading to the production of SA will be enhanced by the addition of extra-chromosomal copies of endogenous genes modified such that expression is constitutive . Where necessary, these genes will be further modified through site-directed mutagenesis to abolish feedback regulation. Enzymatic activities which lead from SA to phenylalanine will be abolished by removing the underlying endogenous genes. These endogenous genes, in addition to a codon-optimized ''Rhodosporidium toruloides'' gene encoding for a phenylalanine ammonia-lysase (PAL) enzyme, will be re-introduced at an extra-chromosomal locus as will modified genes containing inducible promoter/repressor “switches” allowing us to direct metabolism toward CA at will. To further enhance the yield of our desired products, two additional categories of changes will be made to our host strain: (1) Enzymatic activities which direct metabolites away from the desired pathway will be abolished by removing the underlying endogenous genes and (2) transporter proteins responsible for re-uptake of SA or CA from the medium will be removed in the same manner. We are confident that this metabolic engineering strategy will enable us to improve product yield at least five-fold over currently reported values [<html><a href="http://www.ncbi.nlm.nih.gov/pubmed/15824922">1</a>, <a href="http://www.ncbi.nlm.nih.gov/pubmed/22752168">2</a></html>].
 +
 +
 +
 +
<html><center>
 +
<p>
 +
<a href="https://static.igem.org/mediawiki/2012e/f/f2/Upcycled_Aromatics_in_silico_Biobricks.txt">
 +
<img src="https://static.igem.org/mediawiki/2012e/9/9e/Pathway.jpg"> </a></p>
 +
(Click image to see <i>in silico</i> BioBricks)
 +
</center></html>
 +
 +
 +
Our genetic switch mechanism will be designed to tightly regulate gene expression. This will be accomplished by using both an inducer module and a repressor module so that strong expression of the regulated gene is only possible when a dual signal is given. One signal will cause an “inducer protein” to bind a cis regulatory element upstream of the promoter, where it can recruit cell machinery to initiate transcription. This signal alone, however, would be insufficient to allow expression. A repressor cis element located between the inducer cis element and the promoter is normally bound by a “repressor protein” which prevents movement of the transcription machinery down the gene. A second signal will cause the “repressor protein” to dissociate from its cis element, thus clearing the path of transcription machinery and allowing expression. The exact details of this regulatory system are still under investigation – the design of this system may become a focus of next year's Alberta North iGEM team.
 +
 +
 +
 +
<html>
 +
<center>
 +
BioBricks to be made:
 +
<p>
 +
<p>
 +
<table class=MsoNormalTable border=0 cellspacing=0 cellpadding=0
 +
style='border-collapse:collapse;mso-padding-alt:0cm 5.4pt 0cm 5.4pt'>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:solid black 1.0pt;
 +
  border-right:none;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Part name<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Gene/Locus<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Source<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:solid black 1.0pt;
 +
  border-left:none;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Note<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutive promoter<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Inducible promoter (<span class=SpellE>cis</span>)<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Inducible promoter (trans)<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Inducible repressor (<span class=SpellE>cis</span>)<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Inducible repressor (trans)<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Ribosome binding site<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border-top:none;border-left:
 +
  solid black 1.0pt;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Transcription terminator<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>n/a<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Synthetic<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border-top:none;border-left:
 +
  none;border-bottom:solid black 1.0pt;border-right:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>General part<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Transketolase</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_3480<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutively <span class=SpellE>upregulated</span><o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>DAHP synthase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1503<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutively <span class=SpellE>upregulated</span>, Thr326Pro
 +
  = feedback insensitive<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>DHQ synthase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_4923<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutively <span class=SpellE>upregulated</span><o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>DHQ <span class=SpellE>dehydratase</span><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0557<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutively <span class=SpellE>upregulated</span><o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border-top:none;border-left:
 +
  solid black 1.0pt;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Shikimate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> dehydrogenase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0037<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border-top:none;border-left:
 +
  none;border-bottom:solid black 1.0pt;border-right:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Constitutively <span class=SpellE>upregulated</span><o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Shikimate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> kinase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_4924<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>3-phosphoshikimate 1-carboxyvinyltransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1411<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Chorismate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> synthase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1471<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Chorismatemutase</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1410<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Aromatic amino acid aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1561<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Aromatic amino acid aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_3083<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border-top:none;border-left:
 +
  solid black 1.0pt;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Histidinol</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>-phosphate aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0995<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border-top:none;border-left:
 +
  none;border-bottom:solid black 1.0pt;border-right:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='mso-yfti-lastrow:yes;height:15.75pt'>
 +
  <td width=212 valign=bottom style='width:158.9pt;border-top:none;border-left:
 +
  solid black 1.0pt;border-bottom:solid black 1.0pt;border-right:none;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Phenylalanine ammonia-<span class=SpellE>lysase</span><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=79 valign=bottom style='width:58.95pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>&nbsp;<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=72 valign=bottom style='width:54.35pt;border:none;border-bottom:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>R. <span class=SpellE>toruloides</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=269 valign=bottom style='width:201.9pt;border-top:none;border-left:
 +
  none;border-bottom:solid black 1.0pt;border-right:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Introduced as inducible</span></p>
 +
  </td>
 +
</tr>
 +
</table>
 +
 +
<center>
 +
Genes to be knocked out:
 +
<p>
 +
<p>
 +
<table class=MsoNormalTable border=0 cellspacing=0 cellpadding=0
 +
style='border-collapse:collapse;mso-padding-alt:0cm 5.4pt 0cm 5.4pt'>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Shikimate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> kinase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_4924<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>3-phosphoshikimate 1-carboxyvinyltransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1411<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Chorismate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> synthase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1471<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Chorismatemutase</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1410<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Aromatic amino acid aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1561<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Aromatic amino acid aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_3083<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Histidinol</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>-phosphate aminotransferase<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0995<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed and reintroduced as inducible<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border-top:solid black 1.0pt;
 +
  border-left:solid black 1.0pt;border-bottom:none;border-right:none;
 +
  mso-border-top-alt:solid black .5pt;mso-border-left-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Pyrroloquinolinequinone</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_3064<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;border:none;border-top:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;
 +
  height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border-top:solid black 1.0pt;
 +
  border-left:none;border-bottom:none;border-right:solid black 1.0pt;
 +
  mso-border-top-alt:solid black .5pt;mso-border-right-alt:solid black 1.0pt;
 +
  background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Anthranilate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> synthase component I<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0413<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Anthranilate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> synthase component II<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_0416<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Para-<span class=SpellE>aminobenzoate</span> synthase subunit I<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>PPS_1911<i><o:p></o:p></i></span></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
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 +
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 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
 +
  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed<o:p></o:p></span></p>
 +
  </td>
 +
</tr>
 +
<tr style='height:15.75pt'>
 +
  <td width=216 valign=bottom style='width:162.0pt;border:none;border-left:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>Shikimate</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> transporter<o:p></o:p></span></p>
 +
  </td>
 +
  <td width=68 valign=bottom style='width:50.95pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
 +
  mso-fareast-font-family:"Times New Roman";color:black'>shiA</span></span><i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></i></p>
 +
  </td>
 +
  <td width=74 valign=bottom style='width:55.25pt;background:white;padding:
 +
  0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><i><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:
 +
  "Times New Roman";color:black'>P. <span class=SpellE>putida</span></span></i><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'><o:p></o:p></span></p>
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  </td>
 +
  <td width=275 valign=bottom style='width:205.9pt;border:none;border-right:
 +
  solid black 1.0pt;background:white;padding:0cm 5.4pt 0cm 5.4pt;height:15.75pt'>
 +
  <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;mso-line-height-alt:
 +
  5.0pt'><span lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'>Removed<o:p></o:p></span></p>
 +
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 +
</tr>
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<tr style='mso-yfti-lastrow:yes;height:15.75pt'>
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  5.0pt'><span class=SpellE><span lang=EN-US style='font-size:10.0pt;
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  mso-fareast-font-family:"Times New Roman";color:black'>Cinnamic</span></span><span
 +
  lang=EN-US style='font-size:10.0pt;mso-fareast-font-family:"Times New Roman";
 +
  color:black'> acid transporter<o:p></o:p></span></p>
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Latest revision as of 01:28, 27 October 2012

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Genetics

We plan to use a single metabolic pathway for the conversion of glucose to aromatic chemicals. We are currently engineering a Pseudomonas putida strain containing a “switch” system enabling the manufacture of either shikimic acid (SA) or cinnamic acid (CA) as a major product. Future embodiments of this strain will contain additional directing mechanisms whereby a multitude of derivative chemicals can be favored.



Our metabolic engineering strategy is based on the invention of Dr. John Frost (US Patent No. 5168056, 6613552). The enzymatic activities leading to the production of SA will be enhanced by the addition of extra-chromosomal copies of endogenous genes modified such that expression is constitutive . Where necessary, these genes will be further modified through site-directed mutagenesis to abolish feedback regulation. Enzymatic activities which lead from SA to phenylalanine will be abolished by removing the underlying endogenous genes. These endogenous genes, in addition to a codon-optimized Rhodosporidium toruloides gene encoding for a phenylalanine ammonia-lysase (PAL) enzyme, will be re-introduced at an extra-chromosomal locus as will modified genes containing inducible promoter/repressor “switches” allowing us to direct metabolism toward CA at will. To further enhance the yield of our desired products, two additional categories of changes will be made to our host strain: (1) Enzymatic activities which direct metabolites away from the desired pathway will be abolished by removing the underlying endogenous genes and (2) transporter proteins responsible for re-uptake of SA or CA from the medium will be removed in the same manner. We are confident that this metabolic engineering strategy will enable us to improve product yield at least five-fold over currently reported values [1, 2].




(Click image to see in silico BioBricks)


Our genetic switch mechanism will be designed to tightly regulate gene expression. This will be accomplished by using both an inducer module and a repressor module so that strong expression of the regulated gene is only possible when a dual signal is given. One signal will cause an “inducer protein” to bind a cis regulatory element upstream of the promoter, where it can recruit cell machinery to initiate transcription. This signal alone, however, would be insufficient to allow expression. A repressor cis element located between the inducer cis element and the promoter is normally bound by a “repressor protein” which prevents movement of the transcription machinery down the gene. A second signal will cause the “repressor protein” to dissociate from its cis element, thus clearing the path of transcription machinery and allowing expression. The exact details of this regulatory system are still under investigation – the design of this system may become a focus of next year's Alberta North iGEM team.


BioBricks to be made:

Part name

Gene/Locus

Source

Note

Constitutive promoter

n/a

Synthetic

General part

Inducible promoter (cis)

n/a

Synthetic

General part

Inducible promoter (trans)

n/a

Synthetic

General part

Inducible repressor (cis)

n/a

Synthetic

General part

Inducible repressor (trans)

n/a

Synthetic

General part

Ribosome binding site

n/a

Synthetic

General part

Transcription terminator

n/a

Synthetic

General part

Transketolase

PPS_3480

P. putida

Constitutively upregulated

DAHP synthase

PPS_1503

P. putida

Constitutively upregulated, Thr326Pro = feedback insensitive

DHQ synthase

PPS_4923

P. putida

Constitutively upregulated

DHQ dehydratase

PPS_0557

P. putida

Constitutively upregulated

Shikimate dehydrogenase

PPS_0037

P. putida

Constitutively upregulated

Shikimate kinase

PPS_4924

P. putida

Removed and reintroduced as inducible

3-phosphoshikimate 1-carboxyvinyltransferase

PPS_1411

P. putida

Removed and reintroduced as inducible

Chorismate synthase

PPS_1471

P. putida

Removed and reintroduced as inducible

Chorismatemutase

PPS_1410

P. putida

Removed and reintroduced as inducible

Aromatic amino acid aminotransferase

PPS_1561

P. putida

Removed and reintroduced as inducible

Aromatic amino acid aminotransferase

PPS_3083

P. putida

Removed and reintroduced as inducible

Histidinol-phosphate aminotransferase

PPS_0995

P. putida

Removed and reintroduced as inducible

Phenylalanine ammonia-lysase

 

R. toruloides

Introduced as inducible
Genes to be knocked out:

Shikimate kinase

PPS_4924

P. putida

Removed and reintroduced as inducible

3-phosphoshikimate 1-carboxyvinyltransferase

PPS_1411

P. putida

Removed and reintroduced as inducible

Chorismate synthase

PPS_1471

P. putida

Removed and reintroduced as inducible

Chorismatemutase

PPS_1410

P. putida

Removed and reintroduced as inducible

Aromatic amino acid aminotransferase

PPS_1561

P. putida

Removed and reintroduced as inducible

Aromatic amino acid aminotransferase

PPS_3083

P. putida

Removed and reintroduced as inducible

Histidinol-phosphate aminotransferase

PPS_0995

P. putida

Removed and reintroduced as inducible

Pyrroloquinolinequinone

PPS_3064

P. putida

Removed

Anthranilate synthase component I

PPS_0413

P. putida

Removed

Anthranilate synthase component II

PPS_0416

P. putida

Removed

Para-aminobenzoate synthase subunit I

PPS_1911

P. putida

Removed

Shikimate transporter

shiA

P. putida

Removed

Cinnamic acid transporter

 

 

Removed



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Email: ualbertaigem@gmail.com

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