基于流式细胞术的蜘蛛抱蛋属植物倍性检测和核DNA含量测定体系的建立  

Establishment of a System for Estimating Ploidy Level and Nuclear DNA Content of Aspidistra Baed on Flow Cytometry

作　　者：陈婷婷 马均鹏 黄盈萤 梁桂槟 周浩 高乞 CHEN Tingting;MA Junpeng;HUANG Yingying;LIANG Guibing;ZHOU Hao;GAO Qi(School of Marine Sciences and Biotechnology,Guangxi Minzu University,Nanning 530006,China;Nanning Nornal Universisty,Nanning 530001,China)

机构地区：[1]广西民族大学海洋与生物技术学院,南宁530006 [2]南宁师范大学,南宁530001

出　　处：《西北农业学报》2024年第10期1948-1957,共10页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：国家自然科学基金(31560056);广西民族大学引进人才科研启动项目(2018KJQD5)。

摘　　要：为建立适用于蜘蛛抱蛋属植物的倍性检测及其核DNA含量的流式细胞术方法,以蜘蛛抱蛋属植物成熟叶片为材料,比较了WPB解离液、LB01解离液、Galbraith’s解离液以及改良Galbraith’s解离液制备的细胞核悬液的效果。结果表明:改良Galbraith’s解离液能够更广泛地适用于蜘蛛抱蛋属植物成熟叶片的细胞核悬液的制备且效果较好。利用改良Galbraith’s解离液对12种13个居群的蜘蛛抱蛋属植物进行倍性检测,并通过染色体制片计数法进行验证。结果表明:12种蜘蛛抱蛋属植物的DNA含量的荧光强度峰值范围为1753078.81~5817826.99。其中,11种二倍体植物的峰值范围为1753078.81~2937690.80,四倍体辐花蜘蛛抱蛋的峰值为3892503.69,六倍体泰国蜘蛛抱蛋的峰值为5817826.99。此外,以葱为内标,对以上蜘蛛抱蛋属植物的核DNA含量进行测定和估算,首次估测了11种蜘蛛抱蛋植物的核DNA含量,蜘蛛抱蛋的DNA含量与已有报道的结果相近。其中,二倍体植物的核DNA含量为14.16~18.73 Gb;四倍体辐花蜘蛛抱蛋的核DNA含量为28.33 Gb;六倍体泰国蜘蛛抱蛋的核DNA含量为43.03 Gb。结果可为蜘蛛抱蛋属植物的遗传育种研究、全基因组研究以及喀斯特植物的起源与演化研究提供重要科学依据。To establish a flow cytometry method for determining the ploidy level and nuclear DNA content in the genus Aspidistra,we compared the effects of nuclear suspension prepared with WPB buffer,LB01 buffer,Galbraith’s buffer and modified Galbraith’s buffer with the mature leaves.The results showed that the modified Galbraith’s buffer could be more widely and effectively applied to prepare nuclear suspension of mature leaves of Aspidistra.Using the modified Galbraith’s buffer,we detected and verified ploidy level of 12 species from 13 populations within this genus using chromosome counting method.The peak fluorescence intensity of DNA content of 12 species ranged from 1753078.81 to 5817826.99.Among these,the peak value of 11 diploid plant ranged from 1753078.81 to 2937690.80,while the tetraploid A.subrotata was 3892503.69,and the hexploid A.sutepensis was 5817826.99.In addition,the nuclear DNA content of the above plants was measured using Allium fistulosum as internal standard.The nuclear DNA content of 11 species of plants was estimated for the first time,and the DNA content of A.elatior was similar to the reported results.The nuclear DNA content of diploid plants ranged from 14.16 to 18.73 Gb,the tetraploid A.subrotata was 28.33 Gb,and the hexaploid A.sutepensis was 43.03 Gb.The results of this study provide an important scientific basis for the research of genetic breeding,whole-genome research,and the exploration of the origin and evolution of karst plants.

关 键 词：蜘蛛抱蛋属 染色体数目 流式细胞术 倍性水平 核DNA含量 

分 类 号：Q943.2[生物学—植物学]