|Budget Amount *help
¥2,200,000 (Direct Cost : ¥2,200,000)
Fiscal Year 1994 : ¥800,000 (Direct Cost : ¥800,000)
Fiscal Year 1993 : ¥1,400,000 (Direct Cost : ¥1,400,000)
Bacillus thuringiensis is well known for the production of insecticidaltoxin. In the most of case, toxin genes in B.thuringiensis have been reported to be located on plasmid(s) of this bacterium. The developement of a novel host-vector system for molecular breeding in B.thuringiensis was attempted by using temperate phage J7W-1, since this phage genome was revealed to be integrated in a plasmid containing toxin gene in strain AF101.
When the distribution of J7W-1 genome was investigated among type strains of B.thuringiensis, a mutant strain of this phage ; the low level phage induction by ethidium bromide treatment, was found in the type strain of subsp.indiana. The comparison of physical maps of these phage genomes between J7W-1 and its mutant suggested that J7W-1 genome possessed the insertive region for foreign DNA at the length of around 13kb. Furthermore, the addition of cat gene to J7W-1 genome was attempted by homologous recombination. After the chimeric plasmid of pUC18 was constructed by intoducing cat gene and a part of J7W-1 DNA,this plasmid was electropolated into strain AF101 possessing J7W-1 prophage. Although J7W-1 phage containing cat gene was prepared by phage induction of the recombinant cell, cat gene on this phage genome was revealed to be deleted during the phage infection to the host cell of subsp.israelensis. This was considered that the restriction might be occurred in the host cell, since the restriction enzyme : AvaII isosizomer, was identified in subsp.israelensis.