RESTRICTION MAP OF THE 125-KILOBASE PLASMID OF BACILLUS-THURINGIENSISSUBSP ISRAELENSIS CARRYING THE GENES THAT ENCODE DELTA-ENDOTOXINS ACTIVE AGAINST MOSQUITO LARVAE

A large plasmid containing all delta-endotoxin genes was isolated from Bacillus thuringiensis subsp, israelensis; restricted by BamHI, EcoRI , HindIII, KpnI, PstI, SacI, and SalI; and cloned as appropriate libra ries in Escherichia coli, The libraries were screened for inserts cont aining recognition sites for BamHI, SacI, and Sail, Each was labeled w ith P-32 and hybridized to Southern blots of gels with fragments gener ated by cleaving the plasmid with several restriction endonucleases, t o align at least two fragments of the relevant enzymes, All nine BamHI fragments and all eight SacI fragments were mapped in two overlapping linkage groups (with total sizes of about 76 and 56 kb, respectively) , The homology observed between some fragments is apparently a consequ ence pf the presence of transposons and repeated insertion sequences. Four delta-endotoxin genes (cryIVB-D and cytA) and two genes for regul atory polypeptides (of 19 and 20 kDa) were localized on a 21-kb stretc h of the plasmid; without cytA, they are placed on a single BamHI frag ment, This convergence enables subcloning of delta-endotoxin genes (ex cluding cryIVA, localized on the other linkage group) as an intact nat ural fragment.