Team:GZHS-United/Results
Result
The first step of our project is genome DNA extraction, lane 1 is the genome DNA of bacillus thuringiensis .Israelensis, lane 2 is the genome DNA of Bacillus sphaericus. According to the gel, the length of the two genomes are correct.
After genome DNA extraction, we used them as template to amplify Cry4Ba and Mtx1 by PCR, and the results of the amplification were shown in figure 2.
After the target genes were amplified, we have constructed our vector by linking target gene into pEASY-BluntE1 through topo cloning, then transferred the expression vector into E. coli BL21 for protein expression and purification. Figure 3 is the expression vector of Cry4Ba and the outcome after purification. According to the gel, lane 1 is our negative control containing bacterial protein of E. coli BL21, lane 2 is our target protein before purification containing bacterial protein of Cry4Ba E.coli BL21, lane 3 is the protein Cry4Ba after purification. It can be seen from the graph that we have successfully obtained the target protein with high purity.
To produce target protein in a large scale, we tried different induction time and IPTG concentration to find the optimum condition for Cry4Ba expression. It can be seen from figure 4 and 5 that the expressing quantity increased as the increasing of induction time, but the protein expression level has no correlation with IPTG concentration. Therefore, concerning the parameters we mentioned above, we choose to incubate for 6 hours with 0.25mM IPTG as our experimental procedure.
