Part:BBa_K2243011
TP901-1 attL_J23119_TP901-1 attR
The constitutive promoter J23119 is between TP901-1 RDF specific sites and will be inverted by the TP901-1 recombinase and its RDF(or their fusion protein), thus leading to the direction of promoter and change of gene expression. This unit can be used as standard reporter of recombinase TP901-1-RDF fusion protein characterization
Usage
We constructed this part to characterize the recombination efficiency of the excisionase Bxb1 gp35-gp47 fusion protein (BBa_K2243013). It consists of a constitutive promoter (BBa_J23119) flanked by attL and attR sites of recombinase Bxb1 gp35. Upon recombination, the orientation of the constitutive promoter change. As a result, expression of downstream sequence is shut down, and upstream sequence is transcribed.
Biology
J23119 is the consensus sequence of a combinatorial constitutive promoter library family. In nature, attL and attR is generated when Mycobacteriophage Bxb1 integrates its DNA at the attB site of the Mycobacterium smegmatis genome using the viral attP site. With RDF Bxb1 gp47, attL and attR can be recognized and undergo recombination. We obtained the promoter, attL and attR sites by oligo synthesis.
Figure 1. The standard testing system used to characterize the recombinases.
Design note
This part is constructed by Gibson Assembly.
Characterize
We plan to use this part to characterize the recombination efficiency of the integrase Bxb1 gp35 (BBa_K2243013). However, the characterization have not done yet.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 96
Illegal NheI site found at 119 - 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 188
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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