基于转录组测序分析象草木质素合成的研究

摘要

采用高通量测序技术Illumlna HiSeq 2000对高木质素的象草品系eg7和低木质素的象草品系eg87 (对照) 茎组织进行转录组比较测序.测序获得了169630902个序列读取片段 (reads), 包含13788439920nt碱基信息.对reads进行序列组装, 获得87641个单基因簇 (unigene), 平均长度580nt.从长度分布、GC含量等方面对unigene进行评估, 数据显示测序质量好, 可信度高.将获得的unigene与Nr、Nt、Swiss-Prot、COG、GO和KEGG数据库进行序列同源性比较和功能分析, 62557个unigene与其他生物的已知基因具有不同程度的同源性, 象草与高粱序列同源性最高.共鉴定出33323个差异表达基因, 其中上调基因9704个 (29.12％), 下调基因23619个 (70.88％);GO分析显示39968个unigene归为54个功能类别, 大量unigene与细胞进程、代谢过程、催化活性等相关;KEGG pathway分析富集得到127条代谢通路, 包括光合作用、betalain生物合成、苯丙烷类代谢、苯丙氨酸代谢等, 苯丙烷类代谢途径差异基因富集程度高、差异基因数目最多, 达285条, 该途径中64条木质素单体合成酶基因表达上调, 79条ClassⅢ型植物过氧化物酶基因表达下调、22条上调.挑选9个差异基因进行qRT-PCR验证, 9个基因的表达趋势与高通量测序结果一致.为象草的分子生物学研究提供了宝贵的基因组数据, 对于了解象草茎生物合成与木质素调控基因挖掘和多用途定向育种具有指导意义.%Elephant grass (Cenchrus purpureus) is an excellent forage crop and a promising lignocelluloses energy crop;its stem characteristics are very important for forage and feedstock quality.In order to underst and the gene expression profile of elephant grass stem in the absence of a reference genome, we performed de novo transcriptome sequencing and compared the transcriptome of stems with a high lignin content plant (HLCP) eg7 and a low lignin content plant (LLCP) eg87 in elephant grass using Illumina HiSeq 2000 platform.A total of 169630902 high qualities reads were obtained, which were then assembled into 87641 unigenes with an average length of 580nt.All of the 87641 unigenes were compared against the non-redundant protein database (Nr), non-redundant nucleotide database (Nt), swisssprot protein sequence database (Swiss-Prot), gene ontology (GO), cluster of orthologous groups (COG) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) databases for functional annotation;a total of 62557 unigenes were annotated.Most homologous sequences were found between elephant grass with sorghum.33323unigenes showed significant differences in expression between eg7 and eg87 (control), 9704 (29.1％ ) genes were up-regulated and 23619 (70.9％ ) were down-regulated.GO and KEGG pathway analysis revealed that the genes related to photosynthesis-antenna proteins, photosynthesis, betalain biosynthesis, phenylpropanoid biosynthesis are genes of interest.9monolignol biosynthesis-related genes and 8 ClassⅢperoxidase genes were selected as c andidate genes.qRT-PCR analysis showed that the expression of 9 selected unigenes were consistent with the transcriptome data.Our study presented comprehensive transcriptomic data and gene function analysis of C.purpureus stem, providing a valuable resource for future studies of plant breeding in the genus Pennisetum and comparative genome analysis for C4 grasses.