基于RNA-Seq技术分析家蚕卵巢培养细胞(BmN)高表达基因  

Analysis of Highly Expressed Genes in Bombyx mori Ovarian Culture Cells( BmN) Based on RNA-Seq Technology

作　　者：何蕾[1] 薛仁宇[1,2] 曹广力[1,2] 胡小龙[1,2] 贡成良[1,2] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123 [2]苏州大学现代丝绸国家工程实验室,江苏苏州215123

出　　处：《蚕业科学》2014年第3期409-421,共13页ACTA SERICOLOGICA SINICA

基　　金：国家重点基础研究发展计划"973"项目(No.2012CB114600);国家自然科学基金项目(No.31072085;31272500)

摘　　要：为了更全面了解家蚕卵巢培养细胞(BmN)的转录基因及其涉及的功能,对BmN细胞进行深度RNA测序(RNAseq)。得到的总读段(reads)数为80 968 776条,有效RNA-Seq数据68 672 812条,reads的平均长度为94.85 bp,有效数据达到84.81%,证实测序数据可信。将有效数据比对家蚕mRNA序列,发现BmN细胞中的10 182个基因有不同程度表达。利用qRT-PCR测定随机选择BmN细胞中7个基因的相对表达水平,与RNA-Seq的结果略有差异,但总体趋势表现一致。以家蚕Actin3基因作为参照,计算BmN细胞中表达基因读段倍率变化的log2(Fold_change),将log2(Fold_change)>2的基因定义为高表达基因,共有41个基因在BmN细胞中高表达。选取FPKM值>15且表达水平最高的100个基因进行GO注释,主要涉及细胞组分、分子功能和生物学进程,富集在细胞组分中的基因比率高于富集在分子功能、生物学进程中的基因比率。KEGG分析显示这100个高表达基因分布在能量代谢,转录,翻译,折叠、组装和降解以及信号转导等22条通路上。比较发现BmN细胞与家蚕卵巢组织之间的高表达基因存在差异。To further understand the transcriptional genes and their functions of cultured BmN ceils derived from silkworm （Bombyx mori） ovary, deep RNA sequencing of BraN cell was carried out. 80 968 776 paired-end reads were obtained based on RNA-Seq. After removing low-quality reads, 68 672 812 paired-end reads with an average length of 94. 85 bp were obtained. The ratio of valid data to raw data was 84.81%, confirming that the data were genuine and reliable. 10 182 genes, which were expressed at various degrees in BmN cells, were obtained after blasting the valid data to mRNA se- quences of silkworm. The qRT-PCR detection results of 7 randomly selected genes which were expressed in BmN cells were in line with high-throughput RNA sequencing results, also indicating that the RNA-Seq results in this study were credi- ble. The Iog2（Fold _change） of genes relative to the constitutively expressed silkworm Actin3 gene were calculated, and the gene whose log2（Fold change） was more than 2 was defined as highly expressed gene. The results showed that 41genes were highly expressed in BmN cells. GO analysis of the top 100 highly expressed genes with FPKM value 〉15 indicated that these genes were mainly involved in cellular component, molecular function and biological process. The percentage of genes enriched in cellular component was higher than that in both molecular function and biologi- cal processes. KEGG analysis showed that these genes were involved in 22 pathways, mainly enriched in energymetabolism, transcription, translation, folding, sorting and degradation, and signal transduction. Moreover, expression differences of genes were found by comparing highly expressed genes in BmN cells with those in ovary.

关 键 词：家蚕 BmN细胞 基因表达 GO注释 KEGG分析 

分 类 号：S881.2[农业科学—特种经济动物饲养]