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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