草莓果实cDNA文库的构建及鉴定

Construction of a cDNA library of straberry fruit

草莓是研究果实发育的模式植物。基因表达谱分析已成为目前分子生物学领域重要的研究方向。草莓cDNA文库构建是研究果实发育基因表达的基础工作。本研究从草莓白果中提取高质量的总RNA，通过MMLV反转录酶的作用，把RNA中的mRNA反转成cDNA第一链，再利用特异引物通过LD-PCR扩增合成双链cDNA。将纯化带有黏性末端的双链cDNA与λTrip I Ex2载体进行连接，最后对重组载体进行E.coli XL1-Blue体外包埋为cDNA原始文库。经过鉴定，原始文库的滴度为1.13×10^-7 cfu/mL，重组率为100％，插入片段多分布在0.2～2.0 kb之间,表明构建的草莓果实cDNA文库质量较高，能够满足后续的分子生物学研究工作的需要，同时也为其他果树相关研究提供借鉴。

Strawberry is a model plant for fruit development research. Gene expression profiling is an important research field of molecular biology. Thus, construction of cDNA library lays foundation on studying fruit development-related gene expression in strawberry. In this study, total RNA was isolated from fruits of strawberry white fruit stage, single strand cDNA was synthesized based on the extracted mRNAs of total RNA using MMLV reverse transcriptase,and double strand cDNAs were synthesized using PCR, then double strand cDNAs were purified and packaged by the phage packaging system and ligated to the blunted Trip Ex2 vector, finally transformed into the recombinant vector of E.coli XL1-Blue.The results showed that the constructed cDNA library is of the titer of 1.13×10^-7 cfu/ml, the insert sizes is between 0.2～2 kb, and recombination rate of the cDNA library reached 100%, indicating that a higher cDNA library of strawberry fruit is successfully constructed, which meets the needs of subsequent molecular biology research, meanwhile provides a foundation for other fruit tree-related studies.