| Research Abstract |
We evaluated the potential of BmNPV as a new vector by producing temperature-sensitive mutants, and performed functional analysis on these mutants and their virus promoters. We isolated temperature-sensitive mutants of BmNPV with defects at helicase and RNA polymerase using a mutation inducer (BrdU). The construction of new vectors using in vitro packaging was possible by combining the replication ability and particle-formation ability of these mutants. We also determined the complete base sequence of Plutella xylostella granulovirus (PxGV) and provided important information for the development of a new vector of baculovirus. We analyzed promotor of iel, which is an early BmNPV expression gene, to efficiently express extrinsic genes and constructed gene expression vectors with strong activities by promoting the expression using insect transformation hormone and introducing various enhancer elements. We also established a system that efficiently expresses protein from a small genome size by utilizing Periplaneta fuliginosa densovirus (PfD NV).
Moreover, we noted the gene transferring ability of non-LTR type retrotransposon as a possible vector for incorporating genes inserted into the cell nucleus by a virus into the genome, analyzed it by using R2Bm, which is specifically present in rDNA, and demonstrated the possibility of gene insertion into the genome by homologous recombination using the TRRT and host repair mechanisms.
Thus, the basis for an efficient gene insertion technique by a combination of mutant viruses, modification of promoters, and retrotransposon was established.
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