Acta Agrestia Sinica

›› 2012, Vol. 20 ›› Issue (2): 348-351.DOI: 10.11733/j.issn.1007-0435.2012.02.024

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Effects of Different Fluorescent Dyes on Staining Alfalfa Protoplast

LI Yu-zhu1, SHI Shang-li1, HE Yan-yu2   

  1. 1. College of Pratacultural Science, Gansu Agricultural University, Key Laboratory of Grassland Ecosystem, Ministry of Education, Pratacultural Engineering Laboratory of Gansu Province, Sino-U.S. Centers for Grazingland Ecosystem Sustainability, Lanzhou, Gansu Province 730070, China;
    2. Instrumental Research & Analysis Center of Gansu Agricultural University, Lanzhou, Gansu Province 730070, China
  • Received:2011-11-16 Revised:2012-02-25 Online:2012-04-15 Published:2012-07-05

不同荧光染料对苜蓿原生质体染色效果的研究

李玉珠1, 师尚礼1, 贺延玉2   

  1. 1. 甘肃农业大学草业学院 草业生态系统教育部重点实验室 甘肃省草业工程实验室 中美草地畜牧业可持续发展研究中心, 甘肃 兰州 730070;
    2. 甘肃农业大学研究测试中心, 甘肃 兰州 730070
  • 通讯作者: 师尚礼,E-mail:shishl@gsau.edu.cn
  • 作者简介:李玉珠(1979- ),女,陕西宁强人,博士研究生,教师,主要从事牧草和草坪草种质资源保护与育种的研究,E-mail:liyz@gsau.edu.cn
  • 基金资助:
    国家牧草产业技术体系专项资金(CARS-35);牧草种质资源保护与利用(NB2130135);甘肃牧区优质饲草生产技术研究与示范(201003023)资助

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Abstract: The effects of five fluorescent dyes (FDA, Rhodamine B, Rhodamine 6G, Acridine Orange and DAPI) on staining protoplasts obtained from callus of Madicago sativa ‘Gannong No. 4’ were investigated in order to test the viability of protoplasts, count nuclei and survey processes of protoplast fusion. Result showed that FDA, Rhodamine B, Rhodamine 6G and Acridine Orange dye and mark protoplasts of alfalfa clearly, which could identify parental protoplasts from fusion clusters. FDA is an optimal dye for testing the viability of protoplasts. Acridine Orange and DAPI can dye specific nucleus of protoplasts, which are then used to count and survey nuclei in cell fusion. Acridine Orange can also test the viability of fusion cells. These studies suggest a basis for exploring cell fusion conditions and culturing fusion cells by identifying heterocaryon and testing their viability.

Key words: Alfalfa, Protoplast, Nucleus, Dye

摘要： 以紫花苜蓿品种甘农4号(Madicago sativa L.‘Gannong No. 4’)愈伤组织酶解的原生质体为材料,采用FDA、罗丹明B、罗丹明6G、吖啶橙和DAPI共5种荧光染料对原生质体进行染色效果比较,以期为原生质体活力测定、细胞核计数及原生质体融合过程的观察提供参考。结果表明:FDA、罗丹明B、罗丹明6G和吖啶橙均可使紫花苜蓿原生质体清晰染色,可用于细胞融合时亲本原生质体的标识。FDA是检测原生质体活力的理想染料;吖啶橙和DAPI可对细胞核进行特异性染色,可用于融合时细胞核的观察和计数,前者还可用于检测融合细胞的活力。通过对异源融合体的鉴别及其活力的测定,为摸索融合条件和融合细胞的培养提供了理论依据。

关键词: 苜蓿, 原生质体, 细胞核, 染色

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