Difference between revisions of "Part:BBa K3017067"

Revision as of 00:26, 21 October 2019

Construct for testing CRISPRi sgRNA cross-talk with non-target DNA

This asRNA characterization construct is to characterize the cross talking by sgRNA mismatch with different FP.

The asRNA characterization constructs all contain a constitutively expressed dCas9, a fluorescent protein RFP, asRNA under pBAD promoter, and sgRNA (GFP). Under the regulation of pBAD promoter, asRNA is transcripted when arabinose is added to the culture medium. The transcription start site of the pBAD promoter has been identified according to Brzozowska et al. (2018), in which the asRNA has been placed on the Transcription Start Site (TSS). Forl sgRNA transcription, weak Anderson promoter BBa_J23115 of strength 0.15 is used. Since the sgRNA transcribed will be targeting for for gfp, suppression effect on the RFP should be non-desirable. This allows us to assess the cross talking by sgRNA mismatch.

Reference

Characterizing Genetic Circuit Components in E. coli towards a Campylobacter jejuni Biosensor

Natalia Brzozowska, Jane Gourlay, Ailish O’Sullivan, Frazer Buchanan, Ross Hannah, Alison Stewart, Hannah Taylor, Reuben Docea, Greig McLay, Ambra Giuliano, James Provan, Katherine Baker, Jumai Abioye, Julien Reboud, Sean Colloms

bioRxiv 290155; doi: https://doi.org/10.1101/290155

Sequence and Features