甘蓝型油菜胞质体大量制备技术的研究  

STUDIES ON BULK PRODUCTION OF CYTOPLASTS FROM HYPOCOTYL OF BRASSICA NAPUS

作　　者：李昌功[1] 孙海鹏[1] 周嫦[1] 杨弘远[1] 

机构地区：[1]武汉大学生命科学学院植物生殖生物学研究室,武汉430072

出　　处：《武汉植物学研究》1998年第1期77-81,共5页Journal of Wuhan Botanical Research

基　　金：湖北省自然科学基金

摘　　要：用Percoll密度梯度离心法和吖院橙（acridineorange，AO）结合光照处理两种方法，均从甘蓝型油菜下胚轴原生质体制备到胞质体、在2个Percoll梯度、30000g（18000r／min）与12C离心60min的条件下，获得了含胞质体80％以上的群体；以含80、100、120mg／LAO的酶液酶解下胚轴原生质体，纯化后分别给以3h、2h、lh光照（光强度：4000IX）可使原生质体几乎不能分裂，但保持5d以上的生活力。由于光照导致AO－DNA复合体的DNA降解，造成核不可逆失活，从而可以认为经AO与光照处理的原生质体是胞质体。Bulk cytoplasts were produced from hypocotyl protoplasts of B. napus bymeans of a discontinuous Percoll density gradient centrifugation or treatment with acridine orange (AO)followed by exposure to light irradiation. More than 80% cytoplastswere obtained in the interphase between WS solution and 5% Percoll in a discontinuoustwo step gradients after 60 min centrafugation at 12 C 30 000 g(18 000 r/min). The protoplasts treated with 80 mg/L AO (14 -- 18 h during maceration) plus 3 h irradiation(4 000 lx), 100 mg/L AO plus 2 h irradiation or 120 mg/L AO plus 1 h irradiation seldom divided in culture,but could survive for more than 5 days. As light induced fragmentation of AO--DNA complex resulting in degradation of DNA,the nucleus inactivatedirreversibly,thus the products may by considered as cytoplasts.

关 键 词：甘蓝型油菜 胞质体 制备 

分 类 号：S565.4[农业科学—作物学] Q785[生物学—分子生物学]