荧光定量PCR检测家蚕核型多角体病毒在其宿主体内的增殖动态

为了研究家蚕核型多角体病毒(Bombyx mori nuclear polyhedrosis virus，BmNPV)在其宿主幼虫体内不同组织中的增殖动态，对敏感性家蚕品种306幼虫进行经口定量滴注病毒。在接种后9个时间点，对中肠、血淋巴和脂肪体进行取样。以BmNPV DNA 聚合酶基因(dnapol)指示病毒拷贝数，同时以家蚕细胞质肌动蛋白A3(actin A3)基因作为参比基因，用荧光定量PCR的方法分别检测各个时间点的中肠、血淋巴和脂肪体中病毒的拷贝数。结果表明经口感染2 h，病毒进入中肠；12 h，病毒已经到达血淋巴和脂肪体；再经过约12 h的潜伏期，病毒在各组织中开始快速增殖，到84 h各组织中病毒增殖达到平台期。

Detection of proliferation of Bombyx mori nucleopolyhedrovirus in its host by fluorescence quantitative PCR

To investigate the proliferation of Bombyx mori nucleopolyhedrovirus (BmNPV) in its host, the 5th instar larvae of the susceptible silkworm strain 306 were orally administered with determinate dose of BmNPV. After oral ingestion, midgut, haemolymph and fat body tissues were collected at different time to extract DNA. Fluorescence quantitative PCR amplification of BmNPV DNA polymerase gene (dnapol)was used to detect copy number of BmNPV, and each sample was normalized using silkworm cytoplasm actin A3. The results showed that BmNPV could be detected in the midgut 2 hours post infection. BmNPV could be detected in the haemolymph and fat body 12 hours post infection. Through a latent period of 12 hours, rapid proliferation of virus was detected in the midgut, haemolymph and fat body. The proliferation of BmNPV entered a stationary phase 84 hours post infection.