Part:BBa_K2265000
MP6 mutator (original)
The biobrick is a plasmid with one operon under control of the promoter pBAD together with the transcription factor gene AraC, and six other genes that disrupt DNA replication: DnaQ926, Dam, SeqA, EmrR, Ugi and Cda1.
Usage and Biology
The DnaQ926 is a dominant negative variant of the E. coli DNA pol III proofreading domain. Dam has a strong mutator effect due to impaired mismatch repair. The SeqA protein negatively regulates the initiation of DNA replication at the origin of replication. A low expression of SeqA reduces the transcription of the MP6 plasmid in absence of arabinose. The EmrR gene codes for a protein that compromises intracellular dNTP pools. Ugi codes for a protein that inhibit Ung, a mutagenesis preventing enzyme, through mimicry of structural and electronic features of uracil-containing DNA. CDA1 codes for a cytidine deaminase that is reported to promote the mutation of both prokaryotic and eukaryotic genomic DNA.
The biobrick is based on the MP6 plasmid from addgene (Addgene plasmid # 69669). The original plasmid was changed in order to make it a biobrick. The change done to the plasmid was removing a Spel restriction site by changing an A to a C (nucleotide number 4971 in the original MP6 plasmid). A biobrick prefix was also added in front of the araC gene and a biobrick suffix after the CDA1 gene.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1144
Illegal BamHI site found at 1413
Illegal BamHI site found at 2305
Illegal XhoI site found at 3889 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 979
Illegal AgeI site found at 1285
Illegal AgeI site found at 1774 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 4562
Illegal SapI site found at 961
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