生物信息学进展05RNA-Seq数据分析.ppt

ERR365991_1.fastq.gz * 生物信息学进展 （5）RNA-Seq数据分析 提示 我在服务器上建立了两个目录：tools和genome 现将FastQC、Trimmomatic、bwa、bowtie、bedtools和samtools放在tools目录中，将拟南芥、果蝇、人类基因组序列放在genome目录中 大家可以共同使用。现在，大家已经练习过数据的下载、程序的下载、编译、索引的建立，每个人就不需要保留自己的备份。 * 测序数据分析流程 * 1.数据预处理 数据文件 FastQ文件 mapping file 原始数据 FastQ文件 2.序列比对 初步结果 3.数据分析 进一步的结果 4. GO analysis ChIP-Seq RNA-Seq ChIA-PET 。。。 Expressed transcripts detection * Microarrays Sequence-based SAGE - serial analysis of gene expression (3’) CAGE - cap analysis of gene expression (5’) MPSS - massively parallel signature sequencing cDNA EST Expression Exon array ... RNA-seq = RNA sequencing RNA-Seq Purpose Quantify the expression levels of RNAs Identify new transcripts/genes Identify fusion genes Identify alternative splicing Identify single nucleotide variations Identify RNA editing * Characteristics of RNA-Seq Higher sensitivity Expression level changes in 10000 times Higher coverage For unknown genes/transcripts Single-base resolution * Variations in RNA-Seq mRNA Total RNA Small RNA, such as miRNA, tRNA Ribosomal profiling Long non-coding RNA * Challenges RNAs consist of small exons that may be separated by large introns Mapping reads to genome is challenging The relative abundance of RNAs vary wildly 105 – 107 orders of magnitude Since RNA sequencing works by random sampling, a small fraction of highly expressed genes may consume the majority of reads Ribosomal and mitochondrial genes RNAs come in a wide range of sizes Small RNAs must be captured separately PolyA selection of large RNAs may result in 3’ end bias RNA is fragile compared to DNA (easily degraded) * From http://bioinformatics.ca//files/public/BiCG_2012_Module7.pdf 调查显示 * 新一代测序技术在国内的应用情况以及测序中遇到的最大难题和挑战是什么呢？ 测序应用 /news/industry/106307.html RNA-Seq实验过程 * A typical RNA-seq experiment. Briefly, long RNAs are first converted into a library of cDNA fragments through either RNA fragmentation or DNA fragmentation . Sequencing adaptors (blue) are subsequently added to each cDNA fra