棉花纤维素合酶复合体蛋白的分离与鉴定

棉花学报 ›› 2013, Vol. 25 ›› Issue (2): 129-134.

棉花纤维素合酶复合体蛋白的分离与鉴定

李先良^1,2，李　傲¹，彭良才¹，夏　涛^1*

1.华中农业大学作物遗传改良国家重点实验室/生物质与生物能源研究中心/植物科学技术学院/生命科学技术学院，武汉430070；2.荆楚理工学院生物工程学院，湖北荆门 448000

收稿日期:2012-07-09 出版日期:2013-03-15 发布日期:2013-03-15
通讯作者: xiatao@mail.hzau.edu.cn
作者简介:李先良（1976-），男，博士研究生，libusher@sina.com
基金资助:
转基因生物新品种培育重大转项（2009ZX08009-119B）；国家重点基础研究发展计划（2010CB134401）和高等学校科技创新引智计划（B08032）

Characterization of the Cellulose Synthase Complex in Cotton

LI Xian-liang^1,2, LI Ao¹, PENG Liang-cai¹, XIA Tao^1*

1. National Key Laboratory of Crop Genetic Improvement and National Centre of Plant Gene Research / Biomass and Bioenergy Research Centre / College of Plant Science and Technology / College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China; 2. College of Bioengineering, Jingchu University of Technology, Jingmen, Hubei 448000, China

Received:2012-07-09 Online:2013-03-15 Published:2013-03-15

摘要/Abstract

摘要： 选用开花后24 d的棉纤维提取原生质膜，用Triton X-100溶解后通过纤维素合酶1（Gossypium hirsutum cellulose synthase1, GhCESA1）抗体进行免疫共沉淀（Co-Immunoprecipitation, Co-IP），以聚丙烯酰胺凝胶电泳（Sodium dodecyl sulfate polyacrylamide gel electrophoresis, SDS-PAGE）分离免疫共沉淀的产物，用液相色谱-质谱联用仪（Liquid chromatography-mass spectrometry, LC-MS）进行鉴定。结果表明，棉纤维纤维素合酶复合体可能存在68种蛋白，其中包括8种纤维素合酶（Cellolose synthse, CESA），它们涵盖了棉纤维初生壁和次生壁2种类型纤维素合酶。这说明在次生壁形成的细胞中存在2种类型CESA。本研究也表明复合体中存在非CESA蛋白。

关键词: 棉纤维; 纤维素合酶; 复合体; 次生壁

Abstract: To identify the components of CSCs, plasma membrane (PM) was extracted from cotton fibers at 24 days post anthesis (DPA) and solubilized by triton X-100, from which CSCs were isolated by co-immunoprecipitation mediated by Gossypium hirsutum cellulose synthase1(GhCESA1). The proteins isolated by co-immunoprecipitation mediated by GhCESA1 were characterized by western-blotting using the antibodies of GhCESA1 and Gossypium hirsutum sucrose synthase (GhSuSy). The results show there are GhCESA1 and GhSuSy proteins, which indicates this method can isolate the components of CSCs. Then, CSCs proteins through SDS-PAGE were subjected to nanoflow liquid chromatography-tandem mass spectrometry (LC-MS). 68 kinds of proteins were identificated, in which there were 8 kinds of CESA proteins. GhCESA3, 5, 6 can take part in synthesizing cellulose in promary walls, and the other CESA proteins can be responsible for formation of cellulose in secondary walls. This study suggests that the CSC machinary may be aided by other proteins in addition to cellulose synthase proteins.

Key words: cotton fibers; cellulose synthase; complex; secondary wall

中图分类号:

S562:Q814

李先良, 李傲, 彭良才, 夏涛. 棉花纤维素合酶复合体蛋白的分离与鉴定[J]. 棉花学报, 2013, 25(2): 129-134.

LI Xian-Liang, LI Ao, PENG Liang-Cai, XIA Tao. Characterization of the Cellulose Synthase Complex in Cotton[J]. Cotton Science, 2013, 25(2): 129-134.

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