[1]王爽,郭兵福,张丽娟,等.农杆菌介导大豆子叶节转化法外植体选择新方法[J].大豆科学,2016,35(05):723-729.[doi:10.11861/j.issn.1000-9841.2016.05.0723]
　WANG Shuang,GUO Bing-fu,ZHANG Li-juan,et al.New Method for the Selection of Explants in the Agrobacterium Mediated Cotyledon Nodes Transformation in Soybean (Glycine max L)[J].Soybean Science,2016,35(05):723-729.[doi:10.11861/j.issn.1000-9841.2016.05.0723]
点击复制

农杆菌介导大豆子叶节转化法外植体选择新方法

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第35卷 期数: 2016年05期 页码: 723-729 栏目: 出版日期: 2016-09-20
Title:
New Method for the Selection of Explants in the Agrobacterium Mediated Cotyledon Nodes Transformation in Soybean (Glycine max L)
作者:
王爽1; 2; 郭兵福2; 张丽娟2; 郭勇2; 金龙国2; 杨慧2; 邱丽娟2
（1.东北农业大学 农学院，黑龙江 哈尔滨 150030； 2.中国农业科学院 作物科学研究所/国家农作物基因资源与遗传改良重大科学工程，北京 100081）
Author(s):
WANG Shuang1; 2;  GUO Bing-fu2;  ZHANG Li-juan2;  GUO Yong2;  JIN Long-guo2;  YANG Hui2;  QIU Li-juan2
（1.College of Agriculture, Northeast Agricultural University, Harbin 150030, China; 2.The National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI)/MOA Key Lab of Soybean Biology(Beijing), Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China）
关键词:
外植体状态; 瞬时表达; 萌发时间
Keywords:
Explant status;  Transient expression;  Germination time
DOI:
10.11861/j.issn.1000-9841.2016.05.0723
文献标志码:
A
摘要:
大豆遗传转化一直是植物转基因领域的难点之一，农杆菌介导的子叶节法是大豆遗传转化最常用的方法，靶细胞与农杆菌的互作是该方法的关键环节，而外植体的状态是影响靶细胞活力进而影响外植体对农杆菌敏感性的重要因素。本研究利用中黄10、Jack和水里站3个品种材料开展转化试验，根据子叶和下胚轴的颜色和形态变化将外植体划分为6种状态，并以GUS基因瞬时表达效率为指标，比较不同状态外植体靶细胞对农杆菌敏感性，旨在鉴定外植体的最佳制备时期。结果表明：随着萌发时间的增加，外植体状态变化与萌发时间和真叶长度呈正相关，其瞬时表达效率差异显著，且3个受体品种外植体最佳瞬时表达状态存在差异，Jack和中黄10品种的最佳时期为HG、HG+CG、CG这3种状态的外植体，水里站品种的最佳时期YL状态的外植体，研究结果为大豆遗传转化体系的标准化提供了理论依据。
Abstract:
The genetic transformation of soybean is one of the difficulties in plant transgenic and Agrobacterium-mediated cotyledonary node transformation method is the most commonly used in soybean genetic transformation. Interaction between target cells and Agrobacterium is the key of this method and explant status is an important factor that influence sensitivity and regeneration ability.In this study, the explant was divided into 6 types according to the color and status of cotyledon and hypocotyl. Transient transformation was carried out with zhonghuang 10, Jack and Shuilizhan and the index of transient expression efficiency of GUS gene was used to explore the best time to prepare explants. The results showed that the explants status were positively correlated with germination time and leaf length and had effect on transient expression efficiency. The best status of different receptors was different. The best periods of Jack and Zhonghuang 10 were HG, HG+CG and CG, the best periods of Shuilizhan was YL. Those cnclusions could provide a reference for the standardization of soybean genetic transformation system.

参考文献/References:

［1］Jams C. 2014年全球生物技术/转基因作物商业化发展态势［J］.中国生物工程杂志, 2015, 35(1): 1-14.(Jams C. Global biotechnology/GM crops commercialization development trend in 2014［J］. China Biotechnology, 2015, 35(1): 1-14.)

［2］Singh R J, Hymowitz T. Soybean genetic resoures and crop improvement［J］. Genome, 1999, 42: 605-616.
［3］于洋, 侯文胜, 韩天富.农杆菌介导大豆遗传转化研究进展［J］.大豆科学, 2010, 29(4): 696-701. (Yu Y, Hou W S, Han T F. Approaches to Agrobacterium-mediated transformation in soybean［J］. Soybean Science, 2010, 29(4): 696-701.)
［4］潘川芝, 李凤, 戴良英. 大豆子叶节离体再生体系优化研究［J］. 湖南农业科学, 2006, 36(5): 31-32. (Pan F Z, Li F, Dai L Y. Regeneration system of soybean cotyledonary nodes［J］. Human Agricultural Sciences, 2006, 36(5): 31-32.)
［5］Paz M M, Martinez J C, Kalving A B, et al. Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation［J］.Plant Cell Reports, 2006, 25(3):206-213.
［6］Xue R G, Xie H F, Zhang B.A multi-needle-assisted transformation of soybean cotyledonary node cells［J］. Biotechnology Left, 2006, 28(19): 1551-1557.
［7］Guo B F, Guo Y, Wang J, et al. Co-treatment with surfactant and sonication significantly improves Agrobacterium-mediated resistant bud formation and transient expression efficiency in soybean［J］. Journal of Integrative Agriculture 2015, 14(7): 1242-1250.
［8］Miroslav O, Milan B, Jozef S. A comparative structural analysis of direct and indirect shoot regeneration of Papaver somniferumL in vitro［J］.Journal of Plant Physiology, 2000, 157(3): 281-289
［9］Jefferson R A. Assaying chimeric genes in plants: The GUS gene fusion sustem［J］. Plant Molecular Biology Reporter, 1987, 5(4): 387-405.
［10］Cheng M, Chang Y F, Olhoft P M, et al. Cells/tissues conditioning for facilitating T.DNA delivery［G］. USA : Bentham Science Publishers, 2011: 77-107.
［11］赵丽辉, 王立军. 大豆幼苗初生维管系统的解剖学研究［J］.吉林农业大学学报, 1988, 20(1): 42-45. (Zhao L H, Wang L J. Anatomical studies on the primary vascular system of seeding of Glyine max (L) Meer［J］. Jouranl of Jilin Agricultural University, 1988, 20(1): 42-45.)
［12］Baker C M, Mu-oz-Fernandez N, Carter C D. Improved shoot development and rooting from mature cotyledons of sunflower［J］. Plant Cell, Tissue and Organ Culture, 1999, 58(1): 39-49.
［13］Chraibi K M B, Castelle J C, Latche A, et al.Enhancement of shoot regeneration potential by liquid medium culture from mature cotyledons of sunflower (Helianthus annuus L)［J］. Plant Cell Reporter, 1992(b), 10(12): 617-620.
［14］Knittel N, Escandon A S, Hahne G. Plant regeneration at high frequency from mature sunflower cotyledons［J］. Plant Science, 1991, 73(91): 219-226.
［15］Pugliesi C, Cecconi F, Mandolfo A, Baroncelli S. Plant regeneration and genetic variability from tissue cultures of sunflower (Helianthus annuus L)［J］. Plant Breeding, 1990, 106(2): 114-12l.
［16］Hinchee M A W, Connor-Ward D V, Newell C A, et al. Production of transgenic soybean plants using Agrobacterium-mediated DNA transfer［J］. Biotechnology, 1988, 6(8): 915-922.
［17］Yang X F, Yu X Q, Zheng Z, et al.A high-efficiency Agrobacteriu tumefaciens mediated transformation system using cotyledonary node as explants in soybean (Glycine max L)［J］.Acta Physiologiae Plantarum, 2016, 38: 60.

备注/Memo

基金项目：转基因新品种培育重大专项（2014ZX08004-001）。第一作者简介：王爽（1990-），女，硕士，主要从事大豆转基因研究。E-mail：675659474@qq.com。通讯作者：邱丽娟（1963-），女，研究员，博导，主要从事大豆基因资源挖掘与利用研究。E-mail： qiulijuan@caas.cn。

更新日期/Last Update: 2016-09-24