Difference between revisions of "Part:BBa K4907138"

Revision as of 20:12, 11 October 2023

I13453-B0034-ccdA-B0015

Biology

ccdA is the gene found within the ccd operon, encoding the antidote protein (CcdA) that protects cells from the toxic effects of CcdB.CcdA protein is easily degraded by Lonprotease.The cell loses the ccdA gene due to the loss of the F plasmid, causing the cell to succumb to the toxicity of CcdB.

Usage and design

To verify the role of CcdA in resisting the CcdB toxin, we designed a composite part: BBa_K4907138 to characterize ccdA. The constructed circuit was transformed into E. coli DH10β, followed by chloramphenicol selection of positive transformants, and confirmation was carried out through colony PCR and sequencing.

Characterization

Agarose gel electrophoresis (AGE)

In the construction of this circuit, colony PCR and gene sequencing were used to verify the correctness of the transformants. At around 689bp, a target band of approximately 750bp was observed (Fig. 2).

Verification of dual-plasmid transformation

To validate the resistance of CcdA to ccdB, we performed a dual-plasmid transformation.

The results show that E. coli DH10β transformed with both toxins and antitoxins also exhibited good growth. This confirmed the killing effect of the toxin CcdB again and the neutralization of antitoxin CcdA.

Sequence and Features