石正人2006-07-252018-06-292006-07-252018-06-292000http://ntur.lib.ntu.edu.tw//handle/246246/12556本論文以斜紋夜蛾核多角體病毒為主 要研究對象，試選殖斜紋夜蛾核多角體病 毒(Spodoptera litura nucleopolyhedrovirus , SpltNPV )對斜紋夜蛾幼蟲致病力高的基 因，並利用基因重組的方法，將此影響寄 主範圍的基因選殖到加州苜蓿夜蛾核多角 體病毒(Autographa californica nucleopolyhedrovirus , AcMNPV)上，以提 高重組病毒對斜紋夜蛾幼蟲的致病力，擴 增AcMNPV 之寄主範圍。首先利用EcoRI 限制酵素切割SpltNPV DNA，建立基因 庫，進而選殖基因片段於AcMNPV 病毒傳 送載體(pFastBacHTa)上，共篩選到13 種含 有不同DNA 片段之傳送載體。利用Bacto- Bac 表現系統，將斜紋夜蛾核多角體病 毒DNA EcoRI 限制酵素片段，隨機選殖到 AcMNPV 上，產生含有SpltNPV DNA 片 段之AcMNPV 重組病毒。本試驗共構築八 種含有不同限制酵素片段之不同重組病 毒，利用包埋前病毒(pre-occluded virus)形 式，餵食斜紋夜蛾一齡幼蟲，進行生物檢 定，發現這八種重組病毒對幼蟲致病力皆 不超過30％，且八種重組病毒與野生型 AcMNPV 及無包涵體對一齡幼蟲之致病 力，均無顯著差異。結果顯示能夠提高 AcMNPV 致病力之基因並未選殖至這八種 重組病毒中。This study was based on the Spodoptera litura nucleopolyhedrovirus (SpltNPV) to clone the gene which could increase pathogenicity to the larva of S. litura. Furthermore, the gene related to the host range was incorporated to Autographa californica nucleopolyhedrovirus (AcMNPV) by means of genetic engineering to generate recombinant virus. The recombinant virus was used to feed S. litura larvae for determining its the pathogenicity. First of all, the genomic library of SpltNPV was established by digesting DNA of SpltNPV with several restriction enzymes such as EcoRI. DNA fragments from established genomic library were cloned to the transfer vector pFastBacHTa of AcMNPV. Thirteen clones of recombinant transfer vector contained different EcoRI fragments of SpltNPV were obtained. These transfer vectors were used to generate recombinant AcMNPV by using Bac-to-Bac expression vector system. There were eight different recombinant viruses obtained. The preoccluded form virus was used to bioassay 1st instar of S. litura. Results revealed that pathogenicity of 8 recombinant viruses were less than 30%, and there were no significant difference between recombinant viruses and wild type AcMNPV. This indicated that all recombinant viruses did not contain host range related gene of SpltNPV.application/pdf47566 bytesapplication/pdfzh-TW國立臺灣大學昆蟲學系暨研究所斜紋夜蛾核多角體病毒加州苜蓿 夜蛾核多角體病毒寄主範圍Spodoptera lituranucleopolyhedrovirusAcMPNVhost rangereporthttp://ntur.lib.ntu.edu.tw/bitstream/246246/12556/1/892313B002032.pdf