烟芽夜蛾囊泡病毒3h毒株3h-161的克隆和表达谱分析
Cloning and expression of 3h-161 gene encoded by Heliothis virescens ascovirus 3h (HvAV-3h)
陈 斌；徐存翊；黎 妮；谭 赟；于 欢；黄国华
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DOI：10.7679/j.issn.2095-1353.2022.066
作者单位：湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128；湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128；湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128；湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128；湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128；湖南农业大学植物病虫害生物学与防控湖南省重点实验室，长沙 410128
中文关键词：烟芽夜蛾囊泡病毒3h毒株（HvAV-3h）；3h-161；原核表达；时相分析
英文关键词：Heliothis virescens ascovirus 3h; 3h-161; prokaryotic expression; phase analysis
中文摘要：
本文旨在明确烟芽夜蛾囊泡病毒3h毒株（HvAV-3h）编码的第161号开放阅读框（3h-161）的序列特征及表达谱，为深入研究其功能奠定基础。【方法】 采用PCR技术克隆3h-161的CDs序列，构建原核表达载体，并制备高效的多克隆抗体；利用RT-PCR技术检测3h-161的转录情况，利用蛋白免疫印迹法检测3H-161的表达情况。【结果】 3h-161编码区长516 bp，共编码172个氨基酸残基，感染甜菜夜蛾Spodoptera exigua 3龄幼虫后12 h开始转录、48 h开始表达；3H-161在囊泡病毒属内的不同毒株（HvAV-3j、HvAV-3i、HvAV-3e、HvAV-3f和HvAV-3g）间高度保守，序列一致性为100%。【结论】 本研究阐明了3h-161的序列特征、获得了3H-161的多克隆抗体，明确了其表达谱，为进一步研究该基因在HvAV-3h感染甜菜夜蛾过程中发挥的作用奠定基础。
英文摘要：
[Objectives] To
clarify the sequence characteristics and expression profile of the 3h-161 (open reading frame 161) gene in Heliothis
virescens ascovirus 3h (HvAV-3h), thereby laying a foundation for
further study of the function of this gene. [Methods] The CDs sequence of 3h-161 was cloned
using PCR technology, a prokaryotic expression plasmid vector constructed and
efficient polyclonal antibody prepared. RT-PCR and Western blotting technology
was then used to detect the transcription and expression of 3H-161. [Results] The coding regions of 3h-161 are 516 bp in length, encoding 172 amino acids, respectively. Transcription
began at 12 h and expression from 48 h in 3rd instar S. exigua larvae
after infection with HvAV-3h. The putative protein sequences of 3h-161 are highly conservative in the Ascovirus and share 100% amino acid
sequence identity with HvAV-3j, HvAV-3i, HvAV-3e, HvAV-3f and HvAV-3g. [Conclusion] The sequence characteristics of 3h-161 were determined, a polyclonal antibody of 3H-161 was obtained, and the
expression profile of this gene was clarified, all of which lay a foundation
for further study of the function of the 3h-161 gene during the
infection of S. exigua by HvAV-3h.