Molecular cloning of a new crystal protein gene cry1Af1 of Bacillus thuringiensis NT0423 from Korean sericultural farms.
A new cry1Ab-type gene encoding the 130 kDa protein of Bacillus thuringiensis NT0423 bipyramidal crystals was cloned, sequenced, and expressed in a crystal-negative B. thuringiensis host. Hybridization experiments revealed that the crystal protein gene is located on a 44 MDa plasmid of B. thuringiensis NT0423. A strong positive signal detected on the 6.6 kb HindIII fragment from B. thuringiensis NT0423 plasmid DNA was cloned and sequenced. The cry1Ab-type gene, designated cry1Af1, consisted of open reading frame of 3453 bp, encoding a protein of 1151 amino acid residues. The polypeptide has the deduced amino acid sequences predicting molecular masses of 130,215 Da. With both Bt I and Br II promoter sequences were found, the B. thuringiensis NT0423 crystal protein gene promoter closely aligned with those of cry1A-type crystal protein gene. When compared with known sequences of other Cry and Cyt proteins, the Cry1Af1 protein showed maximum 93% sequence identity to Cry1Ab protein of B. thuringiensis subsp. kurstaki. The expressed Cry1Af1 protein in a crystal-negative B. thuringiensis host appears to have strong insecticidal activity against lepidopteran larvae (Plutella xylostella). Crystals containing Cry1Af1 were about six times more toxic than the wild-type crystals of B. thuringiensis NT0423.
Kim
H S
HS
Laboratory of Insect Pathology & Genetic Engineering, Division of Applied Biology and Chemistry, College of Agriculture & Life Sciences, Seoul National University, Seodun-dong 103, Kweonsun-gu, Suwon 441-744, Kyonggi-do, Korea. hskim69@hanmail.net
Li
M S
MS
D000383:Agriculture; D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics* / Q000378:metabolism; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D001427:Bacterial Toxins; D003001:Cloning, Molecular; D004731:Endotoxins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D006460:Hemolysin Proteins; D007723:Korea; D007915:Lepidoptera / Q000187:drug effects; D008969:Molecular Sequence Data; D010572:Pest Control, Biological; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis