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University of Colorado, BoulderCU ScholarChemical & Biological Engineering GraduateTheses & Dissertations Chemical & Biological Engineering
Spring 1-1-2011
Metabolic Engineering of Fatty Acid Production inEscherichia coli as a Biorefining PlatformPaul HandkeUniversity of Colorado at Boulder, [email protected]
Follow this and additional works at: http://scholar.colorado.edu/chbe_gradetds
Part of the Chemical Engineering Commons, and the Molecular Biology Commons
This Dissertation is brought to you for free and open access by Chemical & Biological Engineering at CU Scholar. It has been accepted for inclusion inChemical & Biological Engineering Graduate Theses & Dissertations by an authorized administrator of CU Scholar. For more information, pleasecontact [email protected].
Recommended CitationHandke, Paul, "Metabolic Engineering of Fatty Acid Production in Escherichia coli as a Biorefining Platform" (2011). Chemical &Biological Engineering Graduate Theses & Dissertations. Paper 22.
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GeneForward/Reverse
PrimerPrimer Sequence
accD(Up) ForwardGTCCCCAAAGATAAAACTGGCATCGAACCAGGTTCAGACAGAAAGGTCCCTAGTAGCACACGAGGTCTCT
accD(Up) ReverseCGCCTTGCGGGTGGGAGTAATGTTGCTTTTAATTCGTTCAATCCAGCTCATTTATCACCTCCTTACG
fabD(Up) ForwardTGGATTCACCTGGGGCTCCGCGCTGGTTCGTTTCTAGGATAAGGATTAAAACGTAGCACACGAGGTCTCT
fabD(Up) ReverseTCCAACGGTTTGAGAACCCTGTCCAGGGAACACAAATGCAAATTGCGTCATTTATCACCTCCTTACG
oxyR(Up) ForwardCATTGCTATTCTACCTATCGCCATGAACTATCGTGGCGATGGAGGATGGATAGTAGCACACGAGGTCTCT
oxyR(Up) ReverseATGGCGGTGTTCAGCCAATGCCACCAGGTACTCAAGATCACGAATATTCATTTATCACCTCCTTACG
nuoB(Down) ForwardCCCGGAAACGAACAGTATCGCTAATCGTCAACGCTAACCGCGAGGCATTAAGGTAGCACACGAGGTCTCT
nuoB(Down) ReverseGTAACGGTCGTTCTCACCGTTGGGATCTATGCGGGTGAGCGTATAATCCATGGAGGAGATAACGG
nuoK(Down) ForwardTGCTGAGCAATCGTAAAGACGACAGCGCGAAAAGAAAAACGGAGGAGCACGCGTAGCACACGAGGTCTCT
nuoK(Down) ReverseAAGAACGAATAAGATTGCCGCGAGGATCAGTCCATGTTGTAAGGGGATCATGGAGGAGATAACGG
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Rank Gene Up/Down Description
1 oxyR Up Oxidative stress regulator
2 yfdG Up Predicted bactoprenol-linked glucose translocase
3 cysM Down Cysteine synthase
4 ybbM Up Putative metal resistance protein
5 fabD Up Malonyl-CoA-ACP transacylase
6 yfcF Up glutathione S-transferase
7 nuoK Down NADH:ubiquinone oxidoreductase
8 yajG Up Predicted lipoprotein
9 hisP Down Histidine ABC transporter
10 pdxB Up erythronate-4-phosphate dehydrogenase
11 rpmE Down 50S ribosomal subunit protein L31
12 yihX Up a-D-glucose-1-phosphatase
13 yfcE Down phosphodiesterase
14 thrL Down Threonine biosynthesis
15 folC Up bifunctional polypolyglutamate synthetase / dihydrofolate synthetase
16 rffM Down UDP-N-acetyl-D-mannosaminuronic acid transferase
17 nuoB Down NADH:ubiquinone oxidoreductase
18 ybdD Down conserved protein
19 yjhH Down Predicted lyase/synthase
20 gyrB Up DNA gyrase
21 accD Up acetyl-CoA carboxyltransferase
22 yihY Up predicted inner membrane protein
23 ycgY Up predicted protein
24 ompR Up Transcription factor regulating outer membrane porin genes25 yjjX Up XTPase / ITPase; phosphatase
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"
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0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
0 5 10 15 20 25 30
Con
cent
ratio
n (µ
M)
Time (min)
01 Operator
02 Operator
OC50 Opreator
OC36
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0
5
10
15
20
25
30
35
40
45
50
0 5 10 15 20 25 30
Con
cent
ratio
n (µ
M)
Time (min)
CN = 30
CN = 300
CN = 3000
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PlasmidAntibiotic Chloramphenicol Kanamycin Chloramphenicol Kanamycin
pGR8-Lac 1.3 200 µg/mL > 200 µg/mL 200 µg/mL > 200 µg/mLpGR8-Lac 2.2 500 µg/mL > 200 µg/mL 500 µg/mL > 200 µg/mL
0 mM IPTG 1 mM IPTG
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