Difference between revisions of "Part:BBa K5246035"

Revision as of 15:41, 28 September 2024

BL21(DE3) ΔWecA

Introduction

Strain description

Usage and Biology

WecA is Undecaprenyl-phosphate α-N-acetylglucosaminyl transferase initiating the biosynthesis of enterobacterial common antigen (ECA) and O-antigen PS by catalyzing the transfer of N-acetylglucosamine (GlcNAc)-1-phosphate onto undecaprenyl phosphate to form Und-P-P-GlcNAc. WecA is the first glycosyltransferase, and deleting the wecA gene impairs the ECA synthesis pathway in E.coli. Therefore, lipopolysaccharides containing it are not produced. We wanted to disable the ECA pathway to reduce the metabolic burden on the cell. We aimed to optimize polysaccharide production by creating a more efficient strain. BL21(DE3) is an E.coli expression strain suitable for expressing non-toxic heterologous genes. The BL21(DE3) strain has a lambda DE3 prophage that allows for the expression of T7 RNA polymerase with IPTG. This strain is an E. coli B derivative that lacks the lon and OmpT proteases, which help preserve foreign proteins from degradation.

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal XbaI site found at 185
Illegal SpeI site found at 458
Illegal PstI site found at 715
12
INCOMPATIBLE WITH RFC[12]
Illegal SpeI site found at 458
Illegal PstI site found at 715
Illegal NotI site found at 164
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal XbaI site found at 185
Illegal SpeI site found at 458
Illegal PstI site found at 715
25
INCOMPATIBLE WITH RFC[25]
Illegal XbaI site found at 185
Illegal SpeI site found at 458
Illegal PstI site found at 715
Illegal AgeI site found at 323
1000
COMPATIBLE WITH RFC[1000]

Functional Parameters

genotype

F- ompT hsdSB (rB-, mB-) gal dcm (DE3)(KanR) ΔwecA

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 __NOTOC__
 <partinfo>BBa_K5246035 short</partinfo>
 ===Introduction===
@@ Line 9: / Line 8: @@
 ===Usage and Biology===
-WecA is Undecaprenyl-phosphate &#945;-N-acetylglucosaminyl transferase initiating the biosynthesis of enterobacterial common antigen (ECA) and O-antigen PS by catalyzing the transfer of N-acetylglucosamine (GlcNAc)-1-phosphate onto undecaprenyl phosphate to form Und-P-P-GlcNAc. WecA is the first glycosyltransferase, and deleting the wecA gene impairs the ECA synthesis pathway in E.coli, therefore lipopolysaccharides containing it are not produced. We wanted to disable the ECA pathway to reduce the metabolic burden on the cell. We aimed to optimize polysaccharide production by creating a more efficient strain. BL21(DE3) is an E.coli expression strain suitable for the expression of non-toxic heterologous genes. The BL21(DE3) strain has a lambda DE3 prophage that allows for the expression of T7 RNA polymerase with IPTG. This strain is an E. coli B derivative that lacks the lon and OmpT proteases, which help preserve foreign proteins from degradation.
+WecA is Undecaprenyl-phosphate &#945;-N-acetylglucosaminyl transferase initiating the biosynthesis of enterobacterial common antigen (ECA) and O-antigen PS by catalyzing the transfer of N-acetylglucosamine (GlcNAc)-1-phosphate onto undecaprenyl phosphate to form Und-P-P-GlcNAc. WecA is the first glycosyltransferase, and deleting the wecA gene impairs the ECA synthesis pathway in E.coli. Therefore, lipopolysaccharides containing it are not produced. We wanted to disable the ECA pathway to reduce the metabolic burden on the cell. We aimed to optimize polysaccharide production by creating a more efficient strain. BL21(DE3) is an E.coli expression strain suitable for expressing non-toxic heterologous genes. The BL21(DE3) strain has a lambda DE3 prophage that allows for the expression of T7 RNA polymerase with IPTG. This strain is an E. coli B derivative that lacks the lon and OmpT proteases, which help preserve foreign proteins from degradation.
 ===Sequence and Features===