New strategy for isolating novel nematicidal crystal protein genes from Bacillus thuringiensis strain YBT-1518

Appl Environ Microbiol. 2008 Nov;74(22):6997-7001. doi: 10.1128/AEM.01346-08. Epub 2008 Sep 26.

Abstract

We have developed a strategy for isolating cry genes from Bacillus thuringiensis. The key steps are the construction of a DNA library in an acrystalliferous B. thuringiensis host strain and screening for the formation of crystal through optical microscopy observation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses. By this method, three cry genes--cry55Aa1, cry6Aa2, and cry5Ba2--were cloned from rice-shaped crystals, producing B. thuringiensis YBT-1518, which consists of 54- and 45-kDa crystal proteins. cry55Aa1 encoded a 45-kDa protein, cry6Aa2 encoded a 54-kDa protein, and cry5Ba2 remained cryptic in strain YBT-1518, as shown by SDS-PAGE or microscopic observation. Proteins encoded by these three genes are all toxic to the root knot nematode Meloidogyne hapla. The two genes cry55Aa1 and cry6Aa2 were found to be located on a plasmid with a rather small size of 17.7 kb, designated pBMB0228.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bacillus thuringiensis / chemistry
  • Bacillus thuringiensis / cytology
  • Bacillus thuringiensis / genetics*
  • Bacterial Proteins / analysis
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Toxins / analysis
  • Bacterial Toxins / chemistry
  • Bacterial Toxins / genetics*
  • Cloning, Molecular
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / genetics
  • Electrophoresis, Polyacrylamide Gel
  • Gene Library
  • Genes, Bacterial
  • Microscopy
  • Molecular Sequence Data
  • Molecular Weight
  • Plasmids
  • Sequence Analysis, DNA
  • Survival Analysis
  • Tylenchoidea / drug effects*

Substances

  • Bacterial Proteins
  • Bacterial Toxins
  • DNA, Bacterial

Associated data

  • GENBANK/AF499736
  • GENBANK/EU121521
  • GENBANK/EU121522