Part:BBa_K1639012
T7 expression system for pSB1C3
In our project we needed to express two different genes at the same bacteria, but unfortunately because of high repeats of sequences gene synthesizing company couldn't synthesize it. To solve this we just divided our genes into different plasmids and cotransformed it into same bacteria. For coexpression of genes from different plasmids origins have to be compatible, like ColA and ColE1. pSB1C3 possess ColE1 origin of replication. So it's compatible for expression with plasmid containg ColA origin of replication. But it doesn't contain expression machinery like promoters, terminators. We inserted this part to modify pSB1C3 so that it can be used as expression vector.
Usage and Biology
This system allows to clone and express genes in pSB1C3 vector. In our project some of genes especially motility and flagella related proteins can be impaired by additional amino acids. They differ only 1 amino acid form original state. Cloning in pET45-b vector would add some additional amino acids at the beginning and at the end of these protein. To overcome this we designed this cloning device which contains essential elements for protein expression. System consist of T7 promoter-rbs-terminator and BamHI-SalI restriction sites for cloning. System isn't adding any additional amino acids to protein expressed. Only you need is to ligate system into any vector and then clone gene of interest inside the system.
Results of protein expression are listed at BBa_K1639005
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 167
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 58
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 72
| n/a | T7 expression system for pSB1C3 |