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RNA干扰技术已广泛应用于沉默基因表达的研究.本文分析烟草花叶病毒(TMV)基因序列,选择设计编码小分子发卡结构RNA(hpRNA)的cDNA;并根据农杆菌双元载体质粒p2355多克隆位点区限制性酶切位点,两端分别加入XbaI和BamHI酶切位点;分别合成单链DNA复性后插入到p2355上,经PCR、测序验证,表明已成功构建了具有潜在表达烟草花叶病毒siRNA的植物表达载体.图3表1参26
RNAi technology has been widely used in the study of silencing gene expression.In this paper, we analyzed the sequence of tobacco mosaic virus (TMV) gene and designed the cDNA coding for the small hairpin RNA (hpRNA), and based on the polyclonal plasmid p2355 Site restriction enzyme digestion site, XbaI and BamHI restriction enzyme sites were added to both ends respectively; single-stranded DNA was synthesized and inserted into p2355, and verified by PCR and sequencing. The results showed that it had been successfully constructed with the potential expression of tobacco Mosaic virus siRNA plant expression vector