Difference between revisions of "Part:BBa K3885211"

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	Figure 2.  Fluorescent protein expression intensity after the gene circuit is combined with the CRISPR system.		Figure 2.  Fluorescent protein expression intensity after the gene circuit is combined with the CRISPR system.

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Revision as of 07:20, 20 October 2021

P28-tetR

Under the influence of the P28 promoter, the repressor tetR is produced and binding to tetO sites represses downstream gene expression.

Usage and Biology

The P28-tetR is mainly used to express the gene tetR and thus produce the repressor protein，where P28 is regulated by the σ28 factor to initiate expression. Our gene circuit belongs to the transducer part of the project, where we need to convert molecular signals to fluorescent signals. And the gene tetR produces a repressor protein that can inhibit the expression of fluorescent proteins.

Characterization

The repressor protein produced by tetR can inhibit the expression of fluorescent protein, which is an important part of the gene circuit for signal conversion. First, we verified this part. The fluorescence results were obviously different from those of the control group, which proves that the element can play an ideal role. (Fig.1.a)
We then combined the gene circuit with the CRISPR system to verify the cleavage activity of Cas9 protein.

Design Page

The tetR gene can express repressor protein[1], which inhibits the expression of fluorescent proteins. Therefore, we use Cas9 targeted cleavage of tetR to generate a fluorescent signal by eliminating the inhibitory effect and achieve signal conversion. When binding to the P28 promoter, tetR can achieve a better expression effect, so we construct part P28-tetR.

References

Lutz, R., and Bujard, H. (1997) Independent and Tight Regulation of Transcriptional Units in Escherichia coli via the LacR/O, the TetR/O and AraC/I-1-I-2 Regulatory Elements. Nucleic Acids Res.25 (6), 1203−1210.

Sequence and Features