[Site-specific integration of heterologous gene into Bacillus thuringiensis chromosome and its expression].
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.


CONCLUSIONS
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.
Liu
Ping
P
Key Laboratory of Microbial Molecular Biology of Hunan Province, Life Science College of Hunan Normal University, Changsha 410081, China. ping_liu2008@yahoo.com.cn
Xia
Liqiu
L
Hu
Shengbiao
S
Yan
Li
L
Ding
Xuezhi
X
Zhang
Youming
Y
Yu
Ziniu
Z
D001413:Bacillus thuringiensis / Q000166:cytology* / Q000235:genetics* / Q000378:metabolism; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002876:Chromosomes, Bacterial / Q000235:genetics*; D004274:DNA, Recombinant / Q000235:genetics; D004731:Endotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018628:Gene Dosage; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D006460:Hemolysin Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008853:Microscopy; D010957:Plasmids / Q000235:genetics; D013997:Time Factors
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004274:DNA, Recombinant; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis