摘要
Sox(SRY-related HMG-box)基因是一类重要的转录因子,在动物的性别分化与决定、器官的发生等方面具有重要的调控作用。旨在利用RNA干扰技术证实该基因的功能,将EsSox21b-like基因617 bp的片段克隆至含有两个T7启动子的L4440载体上,构建了EsSox21b-like-L4440干扰载体,将该重组载体转化至RNaseⅢ缺陷型的HT115菌株中,经IPTG诱导菌株体内转录,获得了EsSox21b-like正义和反义单链RNA,经过变性、退火形成大量双链RNA(EsSox21b-like-dsRNA),能满足以后应用RNA干扰试验对于EsSox21b-like基因功能的研究。
Sox（SRY-related HMG-box）gene is a kind of important transcription factor, which functions in sex determination, organogenesis, etc. The gonad-specific expression of this novel gene suggested that it might play an important role in the development of gonad. To determine the exact role of EsSox21b-like gene using RNA interference, a EsSox21b-like-L4440 interference vector was successfully constructed for production of large amount of dsRNA. The target sequence of EsSox21b-like gene was inserted into the L4440 vector containing two T7 promoters. Then the recombinant plasmid was transformed into HT115 bacteria, a RNaseⅢdeficient strain. A large amount of EsSox21b-like sense and antisense RNAs was simultaneously transcripted in vivo when induced by IPTG. After denatured and annealed, EsSox21b-like double strand RNAs（dsRNAs）were produced and the yield of EsSox21b-like-dsRNA was enough for future RNA interference experiment on the role of EsSox21b-like gene.
出处
《生物技术通报》
CAS
CSCD
北大核心
2014年第6期134-138,共5页
Biotechnology Bulletin
基金
国家自然科学基金项目(31272655)
国家科技支撑计划项目(2012BAD26B04)
上海市科委基础重点科研项目(11JC1404600)
上海高校水产学一流学科建设项目(B5005120001)
