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Published on: August 20, 2018
[Site-specific integration of heterologous gene into Bacillus thuringiensis chromosome and its expression]
Ping Liu1, Liqiu Xia, Shengbiao Hu
1Key Laboratory of Microbial Molecular Biology of Hunan Province, Life Science College of Hunan Normal University, Changsha 410081, China. ping_liu2008@yahoo.com.cn
Objective:
To efficiently construct resistance gene-free Bacillius thuringiensis engineered strain that can stably express heterologous gene.
Methods:
We amplified the trigger factor gene located in chromosome of XBU001 strain as homologous arms and constructed an integrative plasmid pKTF12 on the basis of plasmid pKSV7, a temperature sensitive plasmid. We also constructed a recombinant strain KCTF12 containing cry1Ac gene in its chromosome via the integrative plasmid pKTF12.
Results:
Site-specific integration of cry1Ac into XBU001 chromosome did not affect its normal growth. The cry1Ac gene could stably express and form bipyramid crystals in KCTF12. When compared with HTX42 harboring a high-copy number plasmid, the recombinant strain KCTF12 has the merit of advanced sporulation and an increase in spore number.
Conclusion:
The Site-specific integration proved to be a good approach to construct resistance gene-free Bacillius thuringiensis engineered strain that can stably express the heterologous gene.
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