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Nucleotide sequence of the gene coding for a 130-kDa mosquitocidal protein of Bacillus thuringiensis israelensis
T Yamamoto1, I A Watkinson, L Kim
1Agricultural Products Department, DuPont Company, Wilmington, DE 19898.
Abstract:
The nucleotide sequence of pVB131 containing the gene coding for a 130-kDa Bacillus thuringiensis israelensis (B.t.isr) mosquitocidal protein was determined. The pVB131 plasmid was constructed by Sekar and Carlton [Gene 33 (1985) 151-158]. Our sequencing revealed only one open reading frame large enough to code for a protein of 130 kDa. The translation start site was determined by sequencing the protein isolated from B.t.isr. The amino acid sequence of the protein was deduced from the nucleotide sequence, and its Mr was determined as 128,505. Immunological and biochemical analyses of B.t.isr mosquitocidal proteins indicated that the 130-kDa protein coded by pVB131 was indeed expressed in B.t.isr. Comparing the peptide sequence of the 130-kDa B.t.isr toxin with the sequences of other B.t. toxins having activities specific to lepidopteran species showed that several domains were highly homologous. This suggests that they are evolutionarily related to each other, and in the evolutionary process the sequences in the homologous domains that are important to the insecticidal activity have been conserved.
Insights
Researchers sequenced the Bacillus thuringiensis israelensis (B.t.isr) pVB131 plasmid, identifying a gene for a 130-kDa mosquitocidal protein. This protein is evolutionarily related to other Bacillus thuringiensis toxins, suggesting conserved insecticidal domains.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Bacillus thuringiensis israelensis (B.t.isr) produces mosquitocidal proteins crucial for vector control.
- The pVB131 plasmid is known to carry genes for B.t.isr toxins.
- Understanding the genetic basis of mosquitocidal activity is vital for developing effective pest control strategies.
Purpose of the Study:
- To determine the nucleotide sequence of the pVB131 plasmid.
- To identify and characterize the gene encoding the 130-kDa B.t.isr mosquitocidal protein.
- To investigate the evolutionary relationship of this toxin with other B.t. toxins.
Main Methods:
- Plasmid DNA sequencing to determine the nucleotide sequence of pVB131.
- Open reading frame (ORF) analysis to identify potential protein-coding genes.
- Protein sequencing to determine the translation start site.
- Amino acid sequence deduction and molecular weight calculation.
- Immunological and biochemical analyses to confirm protein expression.
- Sequence comparison with known B.t. toxin sequences.
Main Results:
- The nucleotide sequence of pVB131 was determined.
- A single open reading frame coding for a 130-kDa protein was identified.
- The deduced amino acid sequence corresponded to a protein with a molecular weight of 128,505 Da.
- Immunological and biochemical data confirmed the expression of the 130-kDa protein in B.t.isr.
- Significant homology was found between the 130-kDa B.t.isr toxin and lepidopteran-specific B.t. toxins in conserved domains.
Conclusions:
- The pVB131 plasmid contains the gene for a 130-kDa mosquitocidal protein expressed in B.t.isr.
- The 130-kDa B.t.isr toxin shares homologous domains with lepidopteran-specific toxins, indicating a common evolutionary origin.
- Conserved domains within these toxins are likely critical for their insecticidal activity.