Part:BBa_K3020001:Design
recA promoter-Respond to sos response and initiate expression of downstream repair proteins
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
We delete the dsbG coding and following sequence next to dsbG reverse primer, making the functional sequence shorter, so that PCR can be more easy and efficient. The length of remained parts is about half of the original part: K362001.
Besides, AhpCp2D1 has a Pst1 site in the sequence. We succesfully mutated the Pst1 site and make Assembly Compatibility better.
Primers For PCR
| AhpCp2_DsbGp_AhpCp_FP | gcttctagag caatcgcttttactggagcaggaa |
| AhpCp_RP | gga ctgcagcggccgctactagta ctatacttcctccgtgttttcgatgaga |
| ahpC promoter(K362001)has a Pst1 site in the sequence. We designed this part AhpCp2D1, and succesfully mutated the Pst1 site (ctgcag->ctacag). | |
|---|---|
| ahpCp_mut_FP | cacctcagcctacaggcaggcactgaa |
| ahpCp_mut_RP | ttcagtgcctgcctgtaggctgaggtg |
PCR Time Protocol
| Temperature(°C) | Time(sec) | Number of Cycles |
|---|---|---|
| 94 | 120 | -- |
| 94 | 15 | 35 |
| 53 | 30 | |
| 72 | 30 | |
| 72 | 300 | -- |
Source
The recA promoter fragment was cloned from the genome of E.coli K12 MG1655 strain as a template.Please note that the recA promoter sequences from different genomes are not consistent.
References
Norman A , Hestbjerg Hansen L , Sørensen, Søren J. Construction of a ColD cda Promoter-Based SOS-Green Fluorescent Protein Whole-Cell Biosensor with Higher Sensitivity toward Genotoxic Compounds than Constructs Based on recA, umuDC, or sulA Promoters[J]. Appl Environ Microbiol, 2005, 71(5):2338-2346.