Part:BBa_K1884002:Design
VP16AD-CRY2
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 660
Illegal BglII site found at 1119
Illegal BamHI site found at 1598 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 544
Illegal AgeI site found at 1273 - 1000INCOMPATIBLE 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