[1]张晓可,於丙军.大豆幼苗根和叶片原生质的分离与纯化[J].大豆科学,2009,28(04):697-701.[doi:10.11861/j.issn.1000-9841.2009.04.0697]
　ZHANG Xiao-ke,YU Bing-jun.Isolation and Purification of Protoplasts in Roots and Leaves of Soybean Seedlings[J].Soybean Science,2009,28(04):697-701.[doi:10.11861/j.issn.1000-9841.2009.04.0697]
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大豆幼苗根和叶片原生质的分离与纯化

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第28卷 期数: 2009年04期 页码: 697-701 栏目: 出版日期: 2009-08-25
Title:
Isolation and Purification of Protoplasts in Roots and Leaves of Soybean Seedlings
文章编号:
1000-9841(2009)06-0697-05
作者:
张晓可1; 於丙军1; 2
1.南京农业大学生命科学学院，江苏 南京 210095，
2.南京农业大学大豆研究所，农业部国家大豆改良中心，江苏 南京 210095
Author(s):
ZHANG Xiao-ke1; YU Bing-jun1; 2
1.College of Life Sciences，Nanjing Agricultural University，Nanjing 210095；
2.Soybean Research Institute，Nanjing Agricultural University；National Center for Soybean Improvement，Ministry of Agriculture，Nanjing 210095，Jiangsu，China
关键词:
大豆; 根; 叶片; 原生质体; 分离
Keywords:
Soybean; Root; Leaf; Protoplast; Isolation
分类号:
S565.1
DOI:
10.11861/j.issn.1000-9841.2009.04.0697
文献标志码:
A
摘要:
研究了大豆幼苗根和叶片原生质体的分离、纯化方法及其影响因素。结果表明：适宜大豆根和叶片原生质体分离的酶种类、浓度分别为CPW-13M {CPW（细胞清洗液）+13%(W/V)甘露醇}+3%纤维素酶（cellulose R-10）+1.1%果胶酶（macerozyme R-10）+1.0%半纤维素酶（hemicellulase）和0.15% CaCl22H2O+9%甘露醇+1% cellulase R-10+0.20% pectolaseY-23，pH 5.8，酶解温度为28℃。在根酶解时间为16 h时，原生质体产量可高达1.46×105个g-1FW，活力达57.8%；叶片酶解时间为4 h时，原生质体产量可高达1.74×106个g-1FW，活力达70.3%。对于根而言，从产量和活力两方面考虑，其原生质体用23%蔗糖和CPW-18M混合后的下沉法纯化效果较好，而叶片用25%蔗糖的上浮法纯化效果较好。
Abstract:
The methods for isolation and purification of protoplast in roots and leaves of soybean seedling were investigated together in this study.The results showed that，the suitable enzyme species and concentrations for isolation of root protoplasts were CPW-13M+13%(W/V) mannotol+3% cellulose R-10+1.1% macerozyme R-10+1.0% hemicellulase，pH5.8；those for leaf were 0.15% CaCl22H2O+9% mannitol+1% cellulase R-10+0.20% pectolaseY-23，pH 5.8，and the temperature was 28℃.The reasonable enzymatic times for isolation of root and leaf protoplasts were 16 and 4 h，respectively；and the higher protoplast yield and viability were accordingly 1.46×105protoplastsg-1?FW and 57.8%，1.74×106protoplastsg-1FW and 70.3%.The better purification for root protoplasts was the sinking method of 23% sucrose mixed with CPW-18M，that for leaf was the rising method of 25% sucrose.

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备注/Memo

基金项目：国家自然科学基金资助项目（30871462）；江苏省自然科学基金青年科技创新人才（学术带头人）资助项目（BK2007525）。

作者简介：张晓可（1982-）男，硕士研究生，研究方向为植物逆境生物学。
通讯作者：於丙军，教授，博士生导师。E-mail:bjyu@njau.edu.cn。

更新日期/Last Update: 2014-09-22