IBA诱导楸树嫩枝扦插不定根发育的转录组分析  

Transcriptome Profiling of IBA-Induced Adventitious Root Formation in Softwood Cuttings of Catalpa bungei 'Yu-1'

作　　者：张恩亮[1] 马玲玲[1] 杨如同[1] 李林芳[1] 汪庆[1] 李亚[1] 王鹏[1] Zhang Enliang;Ma Lingling;Yang Rutong;Li Linfang;Wang Qing;Li Ya;Wang Peng(Institute of Botany, Jiangsu Province and Chinese Academy of Sciences Nanjing 210014)

机构地区：[1]江苏省中国科学院植物研究所,南京210014

出　　处：《林业科学》2018年第5期48-61,共14页Scientia Silvae Sinicae

基　　金：国家自然科学基金项目(31200509);江苏省重点研发计划(现代农业)项目(BE2016384;BE2017372);江苏省林业三新工程(LYKJ[2017]05)

摘　　要：【目的】利用RNA-seq技术对楸树易生根品种‘豫楸1号’的嫩枝扦插生根的4个发育阶段进行转录组测序和分析,为阐明楸树不定根发育的分子机制奠定理论基础。【方法】首先利用浓度为2 g·L-1IBA处理‘豫楸1号’嫩枝,然后分别提取处理后0天(对照)、1天(激活期)、15天(愈伤形成期)的插穗基部的RNA及25天(不定根形成期)和35天(不定根伸长期)的根部的RNA进行转录组测序;接着测序得到raw reads,通过去除接头、重复序列、低质量的序列,得到高质量的clean reads,使用Trinity软件进行拼接获得contigs;随后,利用BLASTx、RSEM、Cytoscape及Me V4.9.0分子生物学软件对测序结果进行注释和差异基因鉴定;最后随机选取15个差异表达的RNA-Seq数据利用q PCR验证基因的表达。【结果】在62 955个unigenes中,31 646(50.26%)个unigenenes获得了注释;差异基因分析发现了11 100个差异基因,其中包括10 200个特异的和900个共同的差异基因;GO富集分析发现‘细胞骨架’仅在激活期显著富集,而‘DNA代谢过程’仅在愈伤组织形成期显著富集;KEGG富集分析显示参与丙烷合成、糖酵解和植物激素代谢等途径的基因对楸树不定根的形成具有重要的作用,并且随着楸树不定根发育进程的发展,参与糖酵解途径的基因数目逐渐减少而参与苯丙素合成的基因数量却在持续增加,表明在IBA刺激后,代谢通路的动态变化响应了不定根的发育进程。【结论】通过对楸树‘豫楸1号’不定根发育的4个阶段的转录组分析,发现细胞分裂素和乙烯间的互作促进楸树愈伤形成而生长素和油菜素内酯间的互作则促进了楸树不定根的伸长。虽然本研究不能完全解释‘豫楸1号’不定根形成的分子机制,但它可以作为进一步探索与这一复杂过程相关的候选途径和基因的有力工具,可为解析楸树不定根发育的分子机制和其他近缘物种的研究提供帮助�【Objective】 To better understand potential mechanisms involved in adventitious root（ AR） formation,we performed transcriptome analysis of softwood cuttings of Catalpa bungei ‘Yu-1 ’at four stages of AR formation using the Illumina sequencing method.【Method】 After excision,the bases of the cuttings were dipped for 60 s in a solution with 2 g·L^-1 IBA. Samples were harvested at 0（ p0,control）,1（ p1,activation）,15（ p2,callus formation）,25（ p3,root formation） and 35（ p4,root elongation） days after cutting. Total RNAs were extracted and constructed five paired-end libraries. Libraries were sequenced on an Illumina Hi Seq 2000 instrument. Raw reads were cleaned by removing adaptor sequences,empty reads,and low-quality sequences. Clean reads were assembled into non-redundant transcripts using Trinity software. Functional annotation and identification of differentially expressed genes were performed using BLASTx,RSEM,Cytoscape and Me V4. 9. 0. Fifteen differentially expressed unigenes were randomly selected for q PCR validation of our RNA-seq data.【Result】 Following de novo assembly,62 955 unigenes were obtained,31 646（ 50. 26%） of which were annotated. A total of 11 100 differentially expressed genes（ DEGs）,including 10 200 unique and 900 common,were identified in four comparisons. Based on the all GO enrichment networks,cytoskeleton was only significantly enriched in the activation period,while DNA metabolic process was only significantly enriched in the callus formation.Functional annotation analysis revealed that many of these genes were involved in phenylpropanoid biosynthesis,glycolysis,and plant hormone metabolism,suggesting potential contributions to AR formation. Interestingly,the number of DEGs involved in glycolysis decreased while the number of DEGs involved in phenylpropanoid biosynthesis increased following the AR formative process. These results indicated that pathways experienced a dynamic change upon hormone stimulus to occur the corresponding AR formation

关 键 词：楸树 不定根发育 转录组 富集分析 激素 差异表达基因 

分 类 号：S718.46[农业科学—林学]