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林业科学 ›› 2015, Vol. 51 ›› Issue (3): 41-48.doi: 10.11707/j.1001-7488.20150306

• 论文与研究报告 • 上一篇    下一篇

基于油脂合成期油桐种仁转录组数据的α-亚麻酸代谢途径解析

陈昊, 谭晓风   

  1. 中南林业科技大学 经济林培育与保护省部共建教育部重点实验室 经济林育种与栽培国家林业局重点实验室 长沙 410004
  • 收稿日期:2014-07-21 修回日期:2014-08-22 出版日期:2015-03-25 发布日期:2015-04-10
  • 通讯作者: 谭晓风
  • 基金资助:

    国家林业公益性行业科研专项重大项目(201204403); 中南林业科技大学引进高层次人才科研启动基金(2013RJ003)。

Identification of α-Linolenic Acid Metabolism Pathway Based on Transcriptome Data of Vernicia fordii Kernels during Tung Oil Synthesis Stage

Chen Hao, Tan Xiaofeng   

  1. Key Laboratory of Cultivation and Protection for Non-Wood Forest Trees of Ministry of Education Key Laboratory of Non-Wood Forest Products of State Forestry Administration Central South University of Forestry and Technology Changsha 410004
  • Received:2014-07-21 Revised:2014-08-22 Online:2015-03-25 Published:2015-04-10

摘要:

【目的】 油桐种仁中产出的桐油经济利用价值极高。桐油中α-桐酸的含量高达70%,然而植物体内α-桐酸代谢通路的研究还未见报道,这对直接筛选油桐α-桐酸代谢通路相关酶基因造成一定的困难。α-亚麻酸作为α-桐酸的同分异构体,其代谢通路的研究则较为深入,能为α-桐酸代谢通路的解析提供参考。因此,本研究期望在油桐种仁转录组数据的基础上,解析油桐的α-亚麻酸代谢途径,为油桐α-桐酸代谢机理的阐明提供理论参考。此外,通过调控这些基因的表达模式以及开发与之紧密连锁的分子标记,可大大加快油桐遗传改良和分子育种的进程。【方法】 采用RNA-Seq技术对油桐种仁3个不同油脂合成期的转录组进行比较,获得大量差异表达的Unigene,并将这些Unigene归类于128个代谢途径。在此基础上,通过GO分类和Pathway富集性分析,解析油桐α-亚麻酸代谢通路并分析通路中相关酶基因在油脂合成期的表达变化规律。【结果】 通过对Ⅰ,Ⅱ,Ⅲ期的油桐种仁RNA测序,共获得长度为200~3 000个核苷酸的非冗余Unigene序列58 439条,其中能够比对到公共数据库中已知基因序列的Unigene共有41 059条,占所有非冗余基因的70.3%。不同长度的非冗余Unigene序列与数据库中序列匹配的效率不同,越长的序列匹配效率越高。序列长度大于2 000 bp的序列匹配效率达到98.28%,而500~1 000 bp和100~500 bp的序列分别只有78.86%和48.99%的匹配效率。3个种仁油脂合成期的转录组数据中共有105个Unigene可被富集于α-亚麻酸代谢途径,占所有非冗余Unigene的0.47%。从3个转录组数据的两两比较中鉴别出一些差异表达Unigene,其中也有一些可被富集于α-亚麻酸代谢途径。通过在KEGG数据库中进行检索后发现,105个Unigene序列分别对应于14个α-亚麻酸代谢途径关键酶基因,这些基因在其他物种中都有同源基因与之对应。通过基因表达模式分析发现,整体上与合成代谢相关的基因在油脂合成期呈现上调的表达模式,而与分解代谢相关的基因则呈现下调的表达模式。【结论】 在油桐种仁转录组数据的基础上,解析油桐α-亚麻酸代谢途径,获得与α-亚麻酸代谢相关的重要酶基因并分析它们在油脂合成期的表达模式,这对后续研究具有重要的启示作用。

关键词: 油桐, 转录组测序, α-亚麻酸代谢, 基因表达模式

Abstract:

【Objective】 Tung oil extracted from tung tree (Vernicia fordii) kernels has extremely high economic value. Tung oil contains α-eleostearic acid up to 70 percent. However, there are some difficulties in screening enzyme genes involved in α-eleostearic acid metabolism of tung tree due to very few studies being carried out on plant α-eleostearic acid metabolism pathway. Studies on metabolism pathway of α-linolenic acid, the isomers of α-eleostearic acid, were more in-depth and provide an example for analyses of metabolism pathway of α-eleostearic acid. This study was aimed to provide a theoretical basis for explaining the mechanism of α-eleostearic acid metabolism of tung tree through identifying α-linolenic acid metabolism pathway based on transcriptome data of tung tree kernels. It was also intended to largely accelerate the processes of genetic improvement and molecular breeding of tung tree through regulating gene expression profiles and developing molecular markers tightly linked to them. 【Method】 This study compared transcriptomes of tung tree kernels of 3 different oil synthesis stages using RNA-Seq technology and subsequently obtained a lot of differentially expressed Unigenes which can be classified into 128 metabolic pathways. Through GO classification and pathway enrichment analysis, the metabolism pathway of α-linolenic acid was identified and the expression profiles of related genes were analyzed. 【Result】 A total of 58 439 non-redundant Unigene with a 200-3 000 nucleotide-length were identified by RNA-Seq of tung tree kernel RNA of stages I-III. Accounting for 70.3 percentage of all non-redundant Unigenes, a total of 41 059 Unigene sequences could be matched to public database. Non-redundant Unigenes with different sequence length possessed different matching efficiency in alignments against sequences obtained from database. The longer the sequence was, the higher the matching efficiency would be. The matching efficiency of sequences with length greater than 2 000 bp was up to 98.28% whereas the matching efficiency of sequences with 500-1 000 bp and 100-500 bp length was decreased to 78.86% and 48.99%, respectively. Accounting for 0.47 percentages of all non-redundant Unigenes, a total of 105 Unigenes in 3 transcriptome data were enriched into α-linolenic acid metabolism pathway. Each pair-wise comparison of 3 transriptome data identified numbers of differentially expressed Unigenes and some of them were involved in α-linolenic acid metabolism pathway. Through alignment against KEGG database, 105 Unigene sequences were found to correspond to fourteen key enzyme genes involved in α-linolenic acid metabolism. These key enzyme genes were observed to have homologous genes in other species. Expression profiles showed that genes related to anabolism were generally up-regulated whereas genes related to catabolism were generally down-regulated during oil synthesis stage. 【Conclusion】 This study elucidated α-linolenic acid metabolism pathway of tung tree based on its kernel transcriptome data. Furthermore, key enzyme genes involved in α-linolenic acid metabolism were identified and their expression profiles were analyzed. The results of this study play important roles in enlightenment of the follow-up studies.

Key words: Vernicia fordii, transcriptome sequencing, α-linolenic acid metabolism, gene expression profiles

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