Part:BBa_K143001

5' Integration sequence for the AmyE locus of B.subtilis

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

The 5' integration sequence can be added to the front of a Biobrick construct and the 3' integration sequence specific for this locus (Part: BBa_K143002) 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 AmyE locus was the first locus used for integration into B.subtilis by Shimotsu and Henner#1 and is still commonly used in vectors such as pDR111#2, pDL#3 and their derivatives. Integration at the AmyE locus removes the ability of B.subtilis to break down starch, which can be assayed with iodine as described by Cutting and Vander-horn#4. The 5' and 3' integration sequences for the AmyE locus were used to integrate the Imperial 2008 iGEM project primary construct into the B.sutbilis chromosome.

Design Notes

The AmyE integration sequence was taken from the vector after comparison by BLAST to the B.subtilis chromosome to identify the homologous sequences. The sequence present in both the host chromosome and the plasmid at the 5' end of the gene is the 5' sequence required for integration

Source

The 5’ integration sequence was taken from the shuttle vector pDR111 which has been used in many studies on B.subtilis, in particular in the studies of transcriptional control#2,#5,#6

References

<biblio>

1. 1 pmid=3019840
2. 2 pmid=14597697
3. 3 Bacillus Genetic Stock Center [www.bgsc.org]
4. 4 Cutting, S M.; Vander-Horn, P B. Genetic analysis. In: Harwood C R, Cutting S M. , editors. Molecular biological methods for Bacillus. Chichester, England: John Wiley & Sons, Ltd.; 1990. pp. 27–74.
5. 5 pmid=15937167
6. 6 pmid=12169614

</biblio>