Cloning and expression of Bacillus thuringiensis cry11 crystal protein gene in Escherichia coli.
The six most toxic Pakistani isolates of Bacillus thuringiensis (SBS Bt-23, 29, 34, 37, 45 and 47), which were previously characterized for their toxicity against larvae of mosquito, Anopheles stephensi, and the presence of cry4 gene, were used for cry11 (cry4D) gene amplification. A 1.9-kb DNA fragment of cry11 gene was PCR-amplified, cloned in expression vector pT7-7, and then used for transformation of E. coli BL21C. The optimum expression was obtained with 1 mM IPTG at 37 degrees C for 3 h. This gene showed different percentage homologies at protein level with scattered mutations in the toxic region. Biotoxicity assay of recombinant protein showed that Cry11 of SBS Bt 45 (DAB Bt 5) was the most toxic protein against third instar larvae of mosquito, A. stephensi, and has potentiality of a bioinsecticide against mosquitoes.
Bukhari
Dil Ara Abbas
DA
School of Biological Sciences, University of the Punjab, New Campus, Lahore, Pakistan.
Shakoori
Abdul Rauf
AR
D000595:Amino Acid Sequence; D000818:Animals; D001413:Bacillus thuringiensis / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000633:toxicity; D001681:Biological Assay; D003001:Cloning, Molecular; D009033:Culicidae / Q000187:drug effects; D004252:DNA Mutational Analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000737:chemistry / Q000235:genetics* / Q000633:toxicity; D004926:Escherichia coli / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D005798:Genes, Bacterial; D016680:Genome, Bacterial / Q000235:genetics; D006460:Hemolysin Proteins / Q000737:chemistry / Q000235:genetics* / Q000633:toxicity; D008969:Molecular Sequence Data; D009032:Mosquito Control; D011994:Recombinant Proteins / Q000633:toxicity; D015183:Restriction Mapping; D016415:Sequence Alignment; D013171:Spores, Bacterial / Q000187:drug effects / Q000378:metabolism; D014170:Transformation, Genetic / Q000187:drug effects
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D011994:Recombinant Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis