中文摘要：

为高效获得与绿僵菌产孢相关的全长cDNA,采用DSN（duplex-specific nuclease）均一化技术与SMART^TM（switching mechanism at 5′end of RNA transcript）建库技术相结合的方法,构建了杀虫真菌金龟子绿僵菌（Metarhizium anisopliae var.acridum）菌株CQMa102产孢时期的均一化全长cDNA文库。经检测,原始文库滴度为2.1×10^6cfu/mL,库容量超过6×10^6。随机挑取100个克隆,PCR方法测得文库重组率大于95%,插入片段平均长度1500bp。小规模测序分析表明,全长基因的比例超过60%。对组成性表达基因3-磷酸甘油醛脱氢酶G3PDH和微管蛋白β-tubulin均一化前后丰度检测表明,其丰度均有大幅度的下降,基本满足节约筛库的要求。

英文摘要：

In order to obtain full-length genes related to sporulation in Metarhizium anisopliae var. acridum CQMa102, a normalized cDNA library enriched in full-length sequences was constructed using DSN （duplex-specific nuclease）-normalization method combined with SMART^TM （switching mechanism at 5′end of RNA transcript） technique. The titer of un-amplified cDNA library was about 2.1×10^6 cfu/mL and contained 6×10^6 independent clones. The average cDNA inserts size was 1 500 bp with a recombination rate of 95%. Sequencing results and bioinformatics analysis of random picked clones showed that the ratio of full-length cDNA was about 60%. The abundance of transcripts G3PDH and β-tubulin decreased obviously compared with non-normalized samples. These results indicated that the normalized full-length cDNA library has been constructed successfully, which is convenient for further studyins the sporulation mechanism in Metarhizium.