作物学报 ›› 2014, Vol. 40 ›› Issue (03): 424-430.doi: 10.3724/SP.J.1006.2014.00424
郭艳萍**,任成杰**,李志伟,王文斌,张仁和,路海东,刘建超,张兴华,薛吉全*,郭东伟*
GUO Yan-Ping**,REN Cheng-Jie**,LI Zhi-Wei,WANG Wen-Bin,ZHANG Ren-He,LU Hai-Dong,LIU Jian-Chao,ZHANG Xing-Hua,XUE Ji-Quan*,GUO Dong-Wei*
摘要:
玉米胚乳是籽粒淀粉形成的主要场所,胚乳细胞的发育对籽粒的器官建成具有重要意义。分离胚乳细胞原生质体,能为胚乳培养、转录组分析等后续研究提供均一实验材料。本文在借鉴前人研究的基础上,通过调整酶的搭配种类、浓度以及质膜稳定剂、渗透压稳定剂的使用,优化了一套有效的玉米籽粒胚乳原生质体分离方法,并进一步运用流式分选技术纯化原生质体。结果表明,以1%纤维素酶、0.5%离析酶、0.5%半纤维素酶,在渗透压调节剂0.7~0.8 mol L-1和质膜稳定剂0.8mol L-1的MS培养液内,30℃消解4 h,能够获得大量完整的粗制原生质体,二乙酸荧光素(FDA)染色表明纯化的原生质体仍能保持90%以上的生活力。流式分选术可将有活力原生质体从粗制原生质体悬浮液中富集、纯化出来。对一些原生质体分离和流式分选时的技术参数和影响因素进行了讨论。
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