Part:BBa_K2933261
RBS b+Linker h+His+Linker a+Sumo+Linker b+BlaB-14+T7 terminator
This part consists of RBS, protein coding sequence(His+Linker a+Sumo+Linker b+BlaB-14) and T7 terminator,and the biological module can be build into E.coli for protein expression. This part can be prefaced with promoters of different strengths and types to regulate expression function.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 298
- 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 298
Illegal NheI site found at 75
Illegal NheI site found at 1152 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 298
Illegal BglII site found at 187
Illegal BglII site found at 958
Illegal BamHI site found at 386 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 298
- 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 298
- 1000COMPATIBLE WITH RFC[1000]
Usage and Biology
BlaB is a chromosome-encoded MBL produced by Elizabethkingia meningoseptica. It has a broad substrate profile and is produced constitutively. C. meningosepticum is an ubiquitous Gramnegative rod bacterium of clinical relevance because it can cause neonatal meningitis, adult septicemia, and nosocomial infections and is resistant to most β-lactams, including carbapenems.
This pathogen's multidrug-resistant phenotype and attendant lack of adequate molecular mechanistic data limit the current approaches for its effective management in hospitals and public health settings.
Molecular cloning
First, we used the vector pGEX-6p-1 to construct our expression plasmid. And then we converted the plasmid constructed to E. coli DH5α to expand the plasmid largely.We then extracted plasmids and performed double enzyme digestion verification.
Figure 1. (a) The PCR result of BlaB-14. (b) The verification results by enzyme digestion.
After verification, it was determined that the construction is successful. We converted the plasmid to E. coli BL21(DE3) for expression and purification.
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