Elucidation of the mechanism of CryIIIA overproduction in a mutagenized strain of Bacillus thuringiensis var. tenebrionis.
NB176 is a Bacillus thuringiensis mutant derived by gamma-irradiation of NB125 Bacillus thuringiensis var. tenebrionis (Krieg). It exhibits two interesting phenotypes: (i) oligosporogeny and (ii) twofold to threefold overproduction of the CryIIIA protein. Southern profiles of the NB176 strain showed an additional copy(s) of the cryIIIA gene located on a 4 kb HindIII fragment, in addition to the expected cryIIIA gene on a 3 kb HindIII fragment. Each cryIIIA gene-bearing HindIII fragment was cloned from NB176. The restriction map of the 3 kb HindIII fragment was identical to that published by Donovan and coworkers. Sequencing of the 4 kb HindIII fragment showed no alterations in the promoter region of the cryIIIA gene but did show replacement of the region immediately following the cryIIIA open reading frame with a sequence encoding a transposase with 50% amino acid homology to that of Tn1000. These findings suggest that the overproduction phenotype of NB176 results from extra copies of the cryIIIA gene produced from a transposition event(s) induced or stabilized by gamma-irradiation. Integration of additional copies of the cryIIIA gene into the native 90 MDa plasmid of the wild-type B. thuringiensis var. tenebrionis strain resulted in strains that made enormous crystals, many possessing greatly enhanced insecticidal activity.
Adams
L F
LF
Novo Nordisk Biotech, Inc., Davis, California 95616.
Mathewes
S
S
O'Hara
P
P
Petersen
A
A
Gürtler
H
H
D000595:Amino Acid Sequence; D001413:Bacillus thuringiensis / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D001427:Bacterial Toxins; D003001:Cloning, Molecular; D004731:Endotoxins / Q000096:biosynthesis / Q000235:genetics*; D005720:Gamma Rays; D015870:Gene Expression; D005798:Genes, Bacterial; D006460:Hemolysin Proteins; D008969:Molecular Sequence Data; D005810:Multigene Family; D016296:Mutagenesis; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D019895:Transposases
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D009713:Nucleotidyltransferases; D019895:Transposases