Difference between revisions of "Part:BBa K1884002:Design"
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===References===
 
===References===
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Hughes R M, Bolger S, Tapadia H, et al. Light-mediated control of DNA transcription in yeast[J]. Methods, 2012, 58(4): 385-391

Latest revision as of 23:01, 19 October 2016


VP16AD-CRY2


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 660
    Illegal BglII site found at 1119
    Illegal BamHI site found at 1598
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 544
    Illegal AgeI site found at 1273
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 896
    Illegal BsaI.rc site found at 305
    Illegal SapI.rc site found at 413


Design Notes

One powerful application of protein-interaction dimerizers(CRY2-CIB1) is to allow inducible control of a 'split' protein, in which two inactive fragments are brought together to reconstitute a functional protein activity. In principle, using this approach with optically controlled dimerizers would confer light-dependent activity to a diverse range of target proteins. We used the CRY2-CIB1 modules to reconstitute split versions of Gal4 transcription factor proteins. Based on the transient nature of the CRY-CIB interaction,the light pulses would allow precise, dose-dependent control of protein activity. We expressed in yeast the split Gal4 partners Gal4BD- CIB1 and Gal4AD-CRY2 together with upstream activating sequence under control of a galactose-inducible promoter. We incubated the yeast in the dark or subjected them to up to twenty blue-light pulses over 4 h. Immunoblot analysis of Snl1 revealed a strong dose dependence of protein expression in response to light.


Source

Part cloned from pRMH120 of Addgene and we gain the pRMH120 from Shenzhen Engineering Laboratory of Marine Algal Biotechnology.

References

Hughes R M, Bolger S, Tapadia H, et al. Light-mediated control of DNA transcription in yeast[J]. Methods, 2012, 58(4): 385-391