Difference between revisions of "Part:BBa K1985009"
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BBa_K1321333 is a regulatory part is made up of the Arabinose-Inducible promoter, pBAD, and its transcriptional inhibitor/activator, AraC. It was used to give increased control over the expression of part BBa_K1985007. | BBa_K1321333 is a regulatory part is made up of the Arabinose-Inducible promoter, pBAD, and its transcriptional inhibitor/activator, AraC. It was used to give increased control over the expression of part BBa_K1985007. | ||
− | The part was used in pSB1A3 rather than pSB1C3 as it was cotransformed with pec86, a cytochrome maturation factor, which was in a chloramphenicol resistant plasmid. | + | The part was used in pSB1A3 rather than pSB1C3 as it was cotransformed with pec86, a cytochrome maturation factor, which was in a chloramphenicol resistant plasmid. Pec86 is a pACYC184 derivative containing the E.coli genes, ccmABCDEFGH, of the aeg operon, expressed from the tet promoter of the plasmid. The genes are essential for maturation of cytochromes c, i.e. for the covalent attachment of the heme to the protein. |
Biology: | Biology: | ||
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[[File:PSB1A3--AraC-pBAD--MamOPTX gel.jpeg||400px|thumb|centre|Figure 1. Agarose gel of the restriction digest of BBa_K1985009 in pSB1A3, with EcoRI and PstI.]] | [[File:PSB1A3--AraC-pBAD--MamOPTX gel.jpeg||400px|thumb|centre|Figure 1. Agarose gel of the restriction digest of BBa_K1985009 in pSB1A3, with EcoRI and PstI.]] | ||
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+ | ===References=== | ||
Revision as of 13:45, 17 October 2016
AraC pBAD mamOPXT
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1205
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1144
Illegal BamHI site found at 2081 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 4856
Illegal AgeI site found at 979
Illegal AgeI site found at 1946
Illegal AgeI site found at 2030
Illegal AgeI site found at 2939
Illegal AgeI site found at 3600
Illegal AgeI site found at 4169 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 2561
Illegal BsaI.rc site found at 1232
Illegal SapI site found at 961
Illegal SapI.rc site found at 4736
This is a composite part of part BBa_K1321333 (Imperial iGem, 2014), BBa_K1985006, BBa_K1985000, BBa_K1985002, BBa_K1985001 a . It was used in pSB1A3.
Usage and Biology
Usage: The mamO gene was used to initiate the formation of magnetite by "nucleating" the crystal particles, allowing further development. It was used in combination with the proposed electron transport complex of mamO, P and X in vivo to form magnetite in vivo.
BBa_K1321333 is a regulatory part is made up of the Arabinose-Inducible promoter, pBAD, and its transcriptional inhibitor/activator, AraC. It was used to give increased control over the expression of part BBa_K1985007.
The part was used in pSB1A3 rather than pSB1C3 as it was cotransformed with pec86, a cytochrome maturation factor, which was in a chloramphenicol resistant plasmid. Pec86 is a pACYC184 derivative containing the E.coli genes, ccmABCDEFGH, of the aeg operon, expressed from the tet promoter of the plasmid. The genes are essential for maturation of cytochromes c, i.e. for the covalent attachment of the heme to the protein.
Biology: For more information on the biology of mamO,P and X see parts BBa_K1985006, BBa_K1985000,BBa_K1985002 and BBa_K1985001 and for more information on the AraC pBAD promoter please see BBa_K1321333.
Validation
The part was first validated with a diagnostic restriction digest using EcoRI and PstI and agarose gel electrophoresis. The expected band sizes from the digest were: 5478 base pairs for the plasmid backbone and 2114 for the insert. A 1kB plus DNA marker was used to verify the sizes of the bands and it was confirmed that the correct plasmid had been produced.