小鼠pMSV-Slfn5-GFP表达质粒构建及基因结构分析
发布时间：2018-12-27 18:32
【摘要】：目的构建小鼠pMSV-Slfn5-GFP基因重组表达质粒及基因结构分析。方法提取小鼠肝脏组织中总RNA,反转录为cDNA。PCR扩增小鼠Slfn5编码序列片段,并与pGEM-T Easy载体连接,连接产物转化到大肠埃希菌DH5α中。挑取阳性克隆提取质粒,经限制性内切酶HpaⅠ和XhoⅠ双酶切鉴定。酶切鉴定正确的质粒送Macrogen USA测序。测序正确的质粒通过HindⅢ和XhoⅠ与pMSV-GFP连接,并命名为pMSV-Slfn5-GFP。通过UCSC(http://genome.ucsc.Edu/)分析小鼠Slfn5及其家族基因组结构,并通过NCBI确定Slfn5的蛋白结构域。结果 Slfn5全长基因序列克隆到表达载体pMSV-GFP中,酶切鉴定片段大小为2.65kb。Slfn5的AAA_4蛋白结构域由phylogeny.fr确定了该结构在Slfn蛋白家族中的保守性。结论成功构建了小鼠pMSV-Slfn5-GFP全长基因表达载体。
[Abstract]:Objective to construct the recombinant expression plasmid of mouse pMSV-Slfn5-GFP gene and analyze the gene structure. Methods Total RNA, was extracted from mouse liver tissue and reversely transcribed into cDNA.PCR to amplify mouse Slfn5 coding sequence, and then ligated with pGEM-T Easy vector. The ligation product was transformed into Escherichia coli DH5 伪. The positive clones were selected and identified by restriction endonuclease Hpa 鈪，
本文编号：2393444
[Abstract]:Objective to construct the recombinant expression plasmid of mouse pMSV-Slfn5-GFP gene and analyze the gene structure. Methods Total RNA, was extracted from mouse liver tissue and reversely transcribed into cDNA.PCR to amplify mouse Slfn5 coding sequence, and then ligated with pGEM-T Easy vector. The ligation product was transformed into Escherichia coli DH5 伪. The positive clones were selected and identified by restriction endonuclease Hpa 鈪，
本文编号：2393444
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