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Revision as of 09:01, 17 September 2008
5 Integration Sequence for the pyrD locus of B. subtilis
Integration sequences allow DNA to be incorporated into the chromosome of a host cell at a specific locus using leading (5') and trailing (3') DNA sequences that are the same as those at a specific locus of the chromosome. The 5' integration sequence can be added to the front of a Biobrick construct and the 3' integration sequence specific for this locus (BBa_K143009) to the rear of the Biobrick construct to allow integration of the Biobrick construct into the chromosome of the gram positive bacterium B.subtilis.
The PyrD gene has been a target for numerous integration vectors, including the shuttle vectors pPyr-Cm (GenBank Accession number AY464558) and pPyr-Kan (GenBank Accession number AY464559) 1.
Integration into the PyrD locus makes the B.subtilis auxotrophs for uracil and transformants require about 40ug/ml to allow for growth. This allows us to assay for integration by growing a replica plate with no supplemented uracil to negatively select for transformants.
References
<biblio>
- 1 pmid=15109830
</biblio>