紫花苜蓿耐苏打盐碱相关基因的转录组学分析

doi:10.11733/j.issn.1007-0435.2019.04.008

草地学报 ›› 2019, Vol. 27 ›› Issue (4): 848-858.DOI: 10.11733/j.issn.1007-0435.2019.04.008

紫花苜蓿耐苏打盐碱相关基因的转录组学分析

李红¹, 李波², 邬婷婷², 杨曌¹, 方志坚², 焦德志², 孙婴宁²

1. 黑龙江省农业科学院畜牧兽医分院, 黑龙江齐齐哈尔 161005;
2. 齐齐哈尔大学生命科学与农林学院, 抗性基因工程与寒地生物多样性保护黑龙江省重点实验室, 黑龙江齐齐哈尔 161006

收稿日期:2018-12-06 修回日期:2019-04-15 出版日期:2019-08-15 发布日期:2019-09-26
通讯作者: 李波
作者简介:李红(1960-),女,山东莱芜人,研究员,主要从事牧草与饲草与饲料作物育种与栽培研究,E-mail:hljlihong@163.com
基金资助:
齐齐哈尔大学研究生创新科研项目（YJSCX2018_022X）；黑龙江省省属高等学校基本科研业务费科研项目（135209267，YSTSXK201886）；黑龙江省应用技术研究与开发计划重大项目（GA15B105-5）；齐齐哈尔市科技计划一般项目（NYGG-201916）资助

Transcriptomic Analysis of Soda Salt-alkaline Tolerance Related Genes in Alfalfa

LI Hong¹, LI Bo², WU Ting-ting², YANG Zhao¹, FANG Zhi-jian², JIAO De-zhi², SUN Ying-ning²

1. Institute of Animal Husbandry and Veterinary of Heilongjiang academy of Agricultural sciences, Qiqihar, Heilongjiang Province 161005, China;
2. College of Agriculture, Forestry and Life Sciences, Qiqihar University, Heilongjiang Provincial Key Laboratory of Resistance Gene Engineering and Protection of Biodiversity in Cold Areas, Qiqihar, Heilongjiang Province 161006, China

Received:2018-12-06 Revised:2019-04-15 Online:2019-08-15 Published:2019-09-26

摘要/Abstract

摘要： 为了揭示紫花苜蓿适应苏打盐碱环境胁迫机制，本试验采用Illumina HiSeq 4000测序技术对紫花苜蓿叶片进行转录组测序并对基因进行功能预测，测序共获得91 853个Unigenes，总碱基数为65 369 474 bp。Unigenes序列长度分布显示，测序质量较好，可信度高，其中，45 540个Unigenes与其它生物的基因有不同程度的同源性，通过GO，COG及KEGG数据库注释，将Unigenes具体定位到次生代谢物的生物合成途径、抗生素合成途径、光合作用途径等。紫花苜蓿叶片苏打盐碱胁迫响应中GH3，MYB，HSF等转录因子均发生不同程度的上调，而EREBP转录因子总体受到抑制，同时候选了4CL，PP2C基因及相关转录因子。从91 853个Unigenes中共检测到10 949个SSR位点，包括6类核苷酸基序，A/T出现频率最高，其次为AG/CT和AAG/CTT。qRT-PCR荧光定量检测5个基因的表达趋势与RNA-Seq分析结果一致，证明RNA-Seq测序的可靠性。本研究通过对紫花苜蓿转录组研究，为优质牧草的分子生物学研究提供数据库来源。

关键词: 紫花苜蓿, 苏打盐碱, RNA-Seq测序, 功能分类, SSR分析, qRT-PCR验证

Abstract: In order to reveal the mechanism of alfalfa adapting to soda saline-alkali stress,Illumina HiSeq 4000 sequencing technology was used to sequence the transcriptome of alfalfa leaves and predict the function of genes. A total of 91 853 unigenes were obtained. The total base number was 65 369 474 bp. The length distribution of unigenes sequence showed that the quality of sequencing was good and the reliability was high. Among them,45 540 unigenes had different degree of homology with other organisms genes. Through GO,COG and KEGG database annotations specifically locate unigenes in biosynthetic pathways of secondary metabolites,antibiotic biosynthetic pathways,photosynthetic pathways and so on. GH3,MYB,HSF and other transcription factors were up-regulated in response to soda-alkali stress in alfalfa leaves,while EREBP transcription factors were inhibited in general. At the same time,4CL,PP2C genes and related transcription factors were candidates. A total of 10949 SSR loci were detected from 91 853 unigenes,including 6 nucleotide motifs. A/T was the most frequent,followed by AG/CT and AG/CTT. Quantitative detection of five genes by qRT-PCR fluorescence was consistent with RNA-Seq analysis,which proved the reliability of RNA-Seq sequencing. This study provides a database source for the molecular biology research of high quality forage by studying the transcriptome of alfalfa.

Key words: Alfalfa, Soda salt-alkaline, RNA-Seq sequencing, Functional classification, SSR analysis, qRT-PCR validation

中图分类号:

S963.22+3.3

李红, 李波, 邬婷婷, 杨曌, 方志坚, 焦德志, 孙婴宁. 紫花苜蓿耐苏打盐碱相关基因的转录组学分析[J]. 草地学报, 2019, 27(4): 848-858.

LI Hong, LI Bo, WU Ting-ting, YANG Zhao, FANG Zhi-jian, JIAO De-zhi, SUN Ying-ning. Transcriptomic Analysis of Soda Salt-alkaline Tolerance Related Genes in Alfalfa[J]. Acta Agrestia Sinica, 2019, 27(4): 848-858.

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紫花苜蓿耐苏打盐碱相关基因的转录组学分析

Transcriptomic Analysis of Soda Salt-alkaline Tolerance Related Genes in Alfalfa

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