Composite

Part:BBa_K515104:Design

Designed by: Atipat Patharagulpong   Group: iGEM11_Imperial_College_London   (2011-09-07)

J23100 promoter - Antiholin


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 276


Design Notes

During the design process we had to follow the following specification:

Our own GMO must not be harmed by this module

This module relied heavily on modelling during the design process. Since plasmid copy numbers and copy numbers in the genome are variable we had to take this into account when designing the toxin and anti-toxin components. We discovered that the promoter of the holin-endolysin plasmid has to be 40-400 times weaker than that of the holin construct in the genome. This ratio should take the variability between the gene copy numbers into account and make it so that there is at least one holin molecule for every anti-holin molecule that is produced.

Source

The construct was assembled by using PCR on BBa_K112808 and then inserted into a pSB1C3 plasmid through an In-Fusion reaction.

References