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将苜蓿银纹夜蛾(AcNPV)极早期基因IEI启动子控制的neo基因表达盒插入p10型转载体pAcEP106中得到pAcPIneo,将它和野生型AcNPV DNA共转染Sf细胞得到neo~+重组病毒.因为neo基因的表达使得neo~+基因的重组病毒可以在G418的存在下复制,而野生型则不能.将共转染得到的上清液以很低的感染复数连续传代,通过G418的选择作用,使重组病毒得到富集,三代以后重组病毒比例可达90%以上,如此之高的重组比例使得重组病毒的筛选不须经空斑纯化只须有限稀释即可得到.利用杆状病毒早期基因启动子表达neo基因为用早期基因表达外源毒素基因作重组病毒杀虫剂打下了基础,同时也建立了一种简便有效的筛选、富集阳性重组病毒的方法.
The neo gene expression cassette controlled by the IEI promoter of the very early gene of Autographa californica (AcNPV) was inserted into the p10 type vector pAcEP106 to obtain pAcPIneo, which was co-transfected with the wild-type AcNPV DNA into Sf cells to obtain neo ~ + recombinant virus. Because neo gene expression allows the neo ~ + gene recombinant virus to replicate in the presence of G418, but not in the wild type, the supernatants from the co-transfection were serially passaged at very low multiplicity of infection and were selected by G418 , So that the recombinant virus is enriched, three generations later the proportion of recombinant viruses up to 90%, so high recombination ratio makes the recombinant virus screening without plaque purification only limited dilution can be obtained using baculovirus early gene The promoter expressed neo gene for early gene expression of exogenous toxin gene as a recombinant virus pesticide to lay the foundation, but also to establish a simple and effective screening, enrichment positive recombinant virus.