恒河猴大脑前额叶cDNA文库的构建
Construction of a cDNA Library of the Prefrontal Cortex of Rhesus Monkey
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摘要: 为筛选出与认知、记忆相关的脑部表达基因及基因家族，利用TRIzol试剂从恒河猴大脑前额叶组织抽提总RNA，再用纯化试剂盒从总RNA中成功纯化出mRNA。按照Stratagene公司的cDNA Synthesis Kit(200401)、ZAP-cDNA Synthesis Kit(200400)和ZAP-cDNA GigapackⅢ Gold Cloning Kit(200450)三个试剂盒的操作说明，构建了恒河猴大脑前额叶组织的cDNA文库。文库总库容为2.0×10 6克隆；绝大多数的cDNA插入片段≥0.5 kb，平均长度≈1.0 kb；cDNA片段与噬菌体载体重组率为97.3%。文库各项指标均达要求，为克隆大脑PFC区表达基因、测定基因编码区序列、揭示具有多种剪切组合模式等提供了可靠资源，也为相关基因表达调控的研究提供了方便。Abstract: Total RNA was extracted from the prefrontal cortex (PFC) of rhesus monkeys with TRIzol reagent. mRNA was further purified from total RNA using a purifying kit for mRNA. According to the manual of cDNA Synthesis Kit (200401), ZAP-cDNA Synthesis Kit (200400)，and ZAP-cDNA GigapackⅢ Gold Cloning Kit (200450) from Stratagene, we constructed a cDNA-library of the PFC of rhesus monkeys. Our aim was to identify brain-function (such as cognition and memory) related genes and gene families. The cDNA library has a capacity of 2.0×10 6 clones, and 97.3% of them are recombinant clones. The average size of the inserted cDNA fragments is 1.0 kb with the majority of inserts larger than 0.5 kb, which meets the requirement of a standard cDNA library. The PFC cDNA library constructed will be useful in cloning and sequencing brain-function related genes, and in dissecting the alternative splicing patterns of genes expressed in the brain. Meanwhile, it will also serve as a useful resource in understanding the expression regulations within the brain.
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