Detection of the mosquitocidal toxin genes encoding Cry11 proteins from Bacillus thuringiensis using a novel PCR-RFLP method.
A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for detection of cry11 genes from Bacillus thuringiensis was established. Based on the analysis of conserved regions of the cry11 genes, 2 oligonucleotide primers were designed to amplify a 1459-bp fragment of the cry11Aa gene, and a 1471-bp of the cry11Ba and cry11Bb genes. The amplification products were digested with restriction endonuclease HinfI. Exotic B. thuringiensis strains and native isolates collected from soils, leaves and stored product dust of Argentina were analyzed to study the distribution of cry11 genes. The PCR-RFLP patterns revealed the detection of cry11 genes in 3 of 64 exotic strains and in 10 of 107 native B. thuringiensis isolates tested. Just the cry11Aa gene subclass was detected among these bacteria. Since the methodology was also developed to detect cry11Ba and cry11Bb genes, an experimental future confirmation will be required. Based on the results obtained, the PCR-RFLP method presented may be a valuable tool for specific detection of the mosquitocidal toxin genes encoding Cry11 proteins from B. thuringiensis.
Sauka
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DH
Area Bioinsumos Microbianos, Instituto de Microbiología y Zoología Agrícola (IMYZA), Instituto Nacional de Tecnología Agropecuaria (INTA), Castelar, Buenos Aires, 1712, Argentina. dsauka@cnia.inta.gov.ar
Monella
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RH
Benintende
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GB
D001413:Bacillus thuringiensis / Q000235:genetics* / Q000302:isolation & purification; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics*; D001427:Bacterial Toxins / Q000235:genetics*; D017124:Conserved Sequence; D004269:DNA, Bacterial / Q000235:genetics; D004731:Endotoxins / Q000235:genetics*; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D013045:Species Specificity
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; C115468:CRY11BB protein, Bacillus thuringiensis; D004269:DNA, Bacterial; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis