【摘要】 采用GLGI技术扩增本实验室构建的Long-SAGE标签库中的κ-酪蛋白基因。使用SMARTTM RACE cDNA Ampliication Kit通过RT-PCR把奶山羊乳腺组织中的mRNA逆转录成cDNA,分别用3’RACE和5’RACE扩增κ-酪蛋白基因的3’端和5’端。扩增产物经TA克隆后转化大肠杆菌,随机挑取阳性克隆后进行测序分析。通过序列比对、拼接,获得全长的κ-酪蛋白基因。应用GLGI技术成功获得了完整的奶山羊乳腺CSN3序列,为CSN3结构分析和功能研究奠定了重要基础。更多还原

【Abstract】 An optimized procedure of GLGI was adopted to amplify κ-casein gene sequence in Long-SAGE tags of dairy goat mammary gland.SMARTTM RACE cDNA Ampliication Kit was used to mRNA of dairy goat mammary gland tissue reverse transcribe to cDNA by RT-PCR.3′ and 5’ cDNA fragments of κ-casein gene were extended by 3’RACE and 5’RACE respectively.Products of PCR were T/A-cloned and transformed to E.coli.Positive colonies were sequenced.Total length κ-casein gene was obtained by sequence contrasting and splicing.Complete CSN3 sequence of dairy goat mammary gland were successfully identified by applying this high-throughput procedure,which established a basis for further structural and functional analysis of CSN3.更多还原