Team:ECUST/Biocide

Biocide

Description

When the concentration of Fe3+ in the environment is high, the hard skeleton of the microenvironment of the iron bacteria will be destroyed and lose its protection. At this time, the added bactericide will have a greater killing effect on the bacteria. The bactericide we selected was cecropin AD, a cationic chimeric antimicrobial peptide obtained from cecropin, consisting of the first 11 amino acid residues of cecropin A and the last 26 amino acid residues of cecropin D. It has strong antibacterial activity against Gram-positive and negative bacteria.

Design

Result

We insert fragment of Cecropin AD into vector pET-28a(Figure 1)

Figure 1. The vector pET-28a Vector is cut by NcoI and BamHI. Sequence of AD was chemically synthesized and amplified by PCR, then ligated with linearized vector by Ezmax.

The plasmid was transformed to E. coli BL21 and cultured at 37 °C for 12 h. positive monoclonal bacteria were cultured and verified by PCR(Figure 2).

Figure 2. 1% Agarose Gel Electrophoresis of PCR, which shows that our vector was successfully constructed.

We verified the expression of Cecropin AD by SDS-PAGE(Figure 3).

Figure 3. The SDS-PAGE of Cecropin AD Lane 1 :before induction Lane 2,3:after induction

To test the effect of cecropin AD, we measured the growth curve of iron bacteria adding with different concentration of cecropin AD.

Reference

1. Scientific Reports volume7, Article number: 40587 (2017) Efficient biosynthesis of a Cecropin A-melittin mutant in Bacillus subtilis WB700 Shengyue Ji