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2025年8月29日 星期五
农业生物技术学报  2019, Vol. 27 Issue (3): 542-550    DOI: 10.3969/j.issn.1674-7968.2019.03.019
  研究资源与技术改进 本期目录 | 过刊浏览 | 高级检索 |
利用流式细胞术鉴定新疆野杏染色体倍性和DNA含量
李雯雯, 刘立强, 帕米尔·艾尼, 王亚楠, 程功, 廖康*
新疆农业大学 特色果树研究中心,乌鲁木齐 830052
Identification of Chromosomal Ploidy and DNA Content in Xinjiang Armeniaca vulgaris by Flow Cytometry
LI Wen-Wen, LIU Li-Qiang, PAMIER Ai-Ni, WANG Ya-Nan, CHENG Gong, LIAO Kang*
Featured Fruit Tree Research Center, Xinjiang Agricultural University, Urumqi 830052, China
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摘要 分布在我国新疆伊犁河谷的野杏(Armeniaca vulgaris)是第三纪暖温带阔叶林的孑遗群落,其分布区域广阔,具有庞大的实生群体和丰富的遗传多样性。为探索适合杏属植物的流式细胞术(flow cytometry, FCM)检测体系,以快速、准确的对杏属植物的倍性和DNA含量进行检测,本研究选取野杏不同供试材料(幼叶, 老叶),通过不同保存方式(新鲜, 冷藏, 液氮保存)处理,并以紫花苜蓿(Medicago sativa)为外标,利用杏流式细胞术检测体系对10种常用的解离液进行筛选与优化,并检测了野杏的细胞倍性和DNA含量。结果表明,新鲜幼嫩的叶片,使用经优化的MgSO4解离液与Lysis解离液(LB01)的混合液(MgSO4-LB01)制备样品进行检测,其变异系数(coefficient of variation, CV)最低,上样速率达150 events/μL。随着试材冷藏时间的增加,样品CV值随之增加。冷藏4 d以内的幼嫩叶片CV值在5%以内,老叶及液氮保存的叶片制备细胞核悬浮液CV值较高,呈现出分辨率差的峰,背景碎片较多。本研究检测得出供试新疆野杏均为二倍体,DNA含量范围为288~321 Mb/C。新鲜或冷藏4 d之内的幼嫩杏叶片是进行流式细胞术检测的理想材料,经优化的MgSO4-LB01混合液是最适杏的解离液,利用流式细胞术可以快速准确地检测杏属植物的倍性及DNA含量。本研究结果为杏属植物基因组学和遗传进化研究提供基础数据。
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关键词 流式细胞术(FCM)野杏倍性DNA含量    
Abstract:The wild apricot (Armeniaca vulgaris) distributed in Yili Valley of Xinjiang, China, is a relic community of the Tertiary warm temperate broad-leaved forest. It has a vast distribution area, large seedling population and abundant genetic diversity. In this study, different materials (young leaves and old leaves) of wild apricot were treated by different preservation methods (fresh, refrigerated, liquid nitrogen preservation). Using Medicago sativa as external standard, the cell ploidy and DNA content of wild apricot were determined by screening and optimizing 10 common dissociation solutions through flow cytometry (FCM). The results showed that the optimum mixture of MgSO4 buffer (MgSO4) and Lysis buffer (LB01) was used to detect the fresh tender leaves, and its coefficient of variation (CV) was the lowest, and the sample rate was 150 events/μL. Aging leaves could not produce a clear main peak. The CV value of samples increased with the increase of cold storage time. The CV value of young leaves within 4 d cold storage was less than 5%. The CV value of the cell nuclear suspension prepared by the old leaves and liquid nitrogen leaves was higher, showing a peak of poor resolution and more background fragments. The results showed that 20 populations of 2 populations of A. vulgaris were diploid and the content of DNA was 288~321 Mb/C. Fresh young leaves or tender leaves in cold storage within 4 days are ideal materials for FCM. The optimized mixture of MgSO4-LB01 was the best dissociation solution for apricot. FCM could be used to detect ploidy and DNA content of apricot plants quickly and accurately. The results of this study provide technical support for ploidy breeding and omics research of apricot plants. The ploidy and DNA content of wild apricot in Xinjiang are preliminarily discussed, which provide important information for the future study of molecular genetics and cytogenetics.
Key wordsFlow cytometry (FCM)    Armeniaca vulgari    Ploidy    DNA content
收稿日期: 2018-08-13     
ZTFLH:  S662.2  
基金资助:国家重点研发计划(No. 2016YFC0501504)、新疆维吾尔自治区园艺学重点学科基金(No. 2016-10758-3)和自治区研究生科研创新项目
通讯作者: *liaokang01@163.com   
引用本文:   
李雯雯, 刘立强, 帕米尔·艾尼, 王亚楠, 程功, 廖康. 利用流式细胞术鉴定新疆野杏染色体倍性和DNA含量[J]. 农业生物技术学报, 2019, 27(3): 542-550.
LI Wen-Wen, LIU Li-Qiang, PAMIER Ai-Ni, WANG Ya-Nan, CHENG Gong, LIAO Kang. Identification of Chromosomal Ploidy and DNA Content in Xinjiang Armeniaca vulgaris by Flow Cytometry. 农业生物技术学报, 2019, 27(3): 542-550.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2019.03.019     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2019/V27/I3/542
 
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