[1]董丽君,刘灵娣,车文利,等.转Na+/H+逆向转运蛋白基因(AlNHX1)大豆纯合系筛选及生理分析[J].大豆科学,2015,34(03):378-383.[doi:10.11861/j.issn.1000-9841.2015.03.0378]
　DONG Li-jun,Liu Ling-di,CHE Wen-li,et al.Screening of Homogeneous Transgenic Lines with Na+/H+?Antiporter Gene (AlNHX1) in Soybean and Its Physiological Analysis[J].Soybean Science,2015,34(03):378-383.[doi:10.11861/j.issn.1000-9841.2015.03.0378]
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转Na+/H+逆向转运蛋白基因(AlNHX1)大豆纯合系筛选及生理分析

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第34卷 期数: 2015年03期 页码: 378-383 栏目: 出版日期: 2015-06-25
Title:
Screening of Homogeneous Transgenic Lines with Na+/H+?Antiporter Gene (AlNHX1) in Soybean and Its Physiological Analysis
作者:
董丽君1; 刘灵娣2; 车文利3; 张书玲1;  杜环1; 高佳佳1; 刘建凤1
1.河北大学 生命科学学院，河北 保定 071002；?
2.河北省农林科学院 经济作物研究所，河北 石家庄 050051;?
3..廊坊市农林科学院 作物研究所,河北 廊坊 065000
Author(s):
DONG Li-jun1;  Liu Ling-di2;  CHE Wen-li3;  ZHANG Shu-ling1;  DU Huan1;  GAO Jia-jia1;  LIU Jian-feng1
1.College of Life Science, Hebei University, Baoding 071002，China;?
2. Institute of Cash Crops, Hebei Academy of Agricultural and Forestry Sciences, Shijiazhuang 050051, China;?
3. Crops Institute of Academy of Agricultural and Forestry Sciences of Langfang, Langfang 065000, China
关键词:
大豆（Giycine max L）; 钠氢逆向转运蛋白基因（AlNHX1）; 转基因植株; 耐盐能力
Keywords:
Soybean (Gossypium hirsutum L);  Na+/H+?antiporter gene (AlNHX1);  Transgenic plant;  Salt-tolerance ability
DOI:
10.11861/j.issn.1000-9841.2015.03.0378
文献标志码:
A
摘要:
为了获得转Na+/H+逆向转运蛋白基因（AlNHX1）大豆纯合株系并验证其盐胁迫条件下生理反应，对转AlNHX1基因T3代材料进行了PCR筛选和RT-PCR 鉴定，并在水培条件下研究了纯合系植株耐盐性、K+/Na+比值、丙二醛（MDA）、过氧化氢（H2O2）含量和相关抗氧化酶活性变化。结果显示经PCR和RT-PCR 鉴定，野生型植株没有出现目的条带，而转基因株系有相应的目的条带，表明AlNHX1基因已经整合到大豆基因组中并正常转录。在盐胁迫条件下，转基因各株系成活率明显高于野生型对照，地上部干重和根部干重较野生型显著增加，达到了显著差异水平（P＜0.05）。此外，在盐胁迫条件下，转基因幼苗能维持根和叶中较高的K+/Na+比值，同时伴随着植株体内MDA和H2O2含量的下降以及超氧化物歧化酶(SOD)和过氧化酶（POD）的增加。本研究结果表明供试外源AlNHX1基因在大豆高代材料中超表达，提高了转基因大豆耐盐能力，可为进一步培育适应盐渍化土壤环境下生长的大豆新品种提供重要的种质资源。
Abstract:
To develop a salt-tolerant soybean cultivar (Glycine max L), a Na+/H+?antiporter gene AlNHX1 from Aeluropus littoralis was successfully expressed in soybean cultivar TF-29. Further, to obtain homogeneous transgenic lines with expression of AlNHX1 gene and to identify the salt-tolerant ability of the transgenic soybean plants, PCR amplification and RT-PCR detection, plant growth performance, K+/Na+?ratio, MDA and H2O2?and the change of antioxidant enzyme activity of transgenic soybean plants were analyzed under hydroponic condition. The results showed that AlNHX1 gene had been integrated into the soybean genome and expressed in transgenic lines．Under 150 mmol?L-1?NaCl salt stress, transgenic lines showed a significant increase in rate of survival, its shoot dry weight and root dry weight reached significant level (P＜0.05 or P＜0.01) compared with wild type plants. The different phenotypes of transgenic soybean and wild type plants were also observed, the transgenics displayed robust performance whereas growth by WT plants was significantly reduced under stress conditions. Also, a higher ratio of K+?to Na+?in the leaves and roots of the transgenic plants was retaining, accompanied with the decline of malondialdehyde (MDA) and hydrogen peroxide (H2O2) and the higher activities of antioxidant enzymes including superoxide dismutase (SOD) and peroxidase (POD) under salt stress.These findings proved that over-expression of exogenous AlNHX1 in soybean have enhanced salt tolerance, and they have significant implications for increasing crop yields on high-salinity soils

参考文献/References:

［1］Launchli A. Salt exclusion: an adaptation of legumes for crops and pastures under saline conditions［M］// Staples RC, Toeniessen G H.Salinity tolerance in plants Strategies for crop improvement. New York:Wiley, 1984:171-187

［2］Maas E V, Hoffman G J.Crop salt tolerance current assessment ［J］.Journal of Irrigation and Drainage Engineering-asce, 1977, 103(2): 115-134
［3］陈观平, 王慧中, 施农农, 等 Na+/H+逆向转运蛋白与植物耐盐性关系研究进展［J］.中国生物工程杂志, 2006, 26(5): 101-106 (Chen G P, Wang H Z, Shi N N et al Research the advance on the relation of Na+/H+?antiporter protein and salt tolerance ［J］.China Biotechnology, 2006, 26(5): 101-106)
［4］Blumwal E. Sodium transport and salt tolerance in plants ［J］Cell Biology, 2000, 12: 431-434
［5］Cao D, Hou W S, Liu W, et al. Overexpression of TANHX2 enhances salt tolerance of ‘composite’ and whole transgenic soybean plants ［J］. Plant Cell Tissue and Organ Culture, 2011, 107(3): 541-552
［6］王敏娟, 侯文胜, 王庆钰, 等. 过表达GmNHX1基因提高大豆根系的耐盐性［J］. 大豆科学, 2011, 30(6): 889-894 (Wang M J, Hou W S, Wang Q Y, et al.Enhancing salt tolerance of soybean roots by overexpression of GmNHX
?［J］. Soybean Science, 2011, 30(6): 889-894)
［7］魏立兴.GmABA 8’OH基因在大豆中的表达及耐盐性分析［D］.长春：东北师范大学, 2012.(Wei L X.The expression of GmABA8’OH gene and the analysis of salt tolerance in soybean ［D］Changchun:Northeast Normal University, 2012)
［8］刘建凤, 崔栗, 安利佳.转AlNHX1基因大豆后代遗传稳定性及耐盐性分析［J］. 河北大学学报, 2012, 32(2): 173-179 (Liu J F, Cui L, An L J. Genetic stability analysis and salt tolerance of the progenies of soybean transformed AlNHX1 gene ［J］ Journal of Hebei University, 2012, 32(2): 173-179
［9］Liu J F, Zhang S L, Dong L J, et al. Incorporation of a Na+/H+?antiporter gene from Aeluropus littoralis confers salt tolerance in soybean ［J］. Indian Journal of Biochemistry and Biophysics, 2014, 51(1): 58-65
［10］吴畏，苏乔，刘建凤, 等.转甜菜碱醛脱氢酶基因玉米的田间筛选及生理分析［J］沈阳农业大学学报，2007, 38(4):612-614 (Wu W, Su Q, Liu J F, et al.Screening in field and physiological analysis on transgenic maize with Betaine aldehyde dehydrogenase gene ［J］ Journal of Shenyang Agricultural University, 2007, 38(4): 612.614)
［11］蒋明义, 郭绍刚.渗透胁迫下稻苗铁催化的膜脂过氧化作用［J］.植物生理学报, 1996, 22(1): 6-12(Jiang M Y, Guo S G. Iron catalyst on membrane lipid peroxidation of rice seedling under osmotic stress［J］.Plant Physiology Journal, 1996, 22(1): 6-12)
［12］汤章城.现代植物生理学实验指南［M］.北京: 科学出版社, 1999.(Tang Z C. Guide the modern plant physiology experiment ［M］. Beijing: Science Press, 1999)
［13］Prochazkova R K D, Sairam G C, Srivasava D V.Single oxidative stress and antioxidant activity as the basis of senescence in maize leaves ［J］Plant Science, 2001, 161(4):765-771
［14］Apse M P, Aharon G S, Snedden W A, et al. Salt tolerance conferred by overexpression of a vacuolar Na+?/ H +?antiporter in Arabidopsis ［J］ Science, 1999, 285(5431): 1256-1258
［15］王艳, 曾幼玲, 贺宾, 等. 农杆菌介导-NHX-基因转化甘蓝型油菜的研究［J］.作物学报, 2006, 32(2): 278-282(Wang Y, Ceng Y L, H B, et al.Research of NHX gene transformation in Brassica napus by Agrobacterium tumefaciens［J］ Acta Agronomica Sinica, 2006, 32(2): 278-282)
［16］Viswanathan C, Andre J, Zhu J K.Understanding and improving salt tolerance in plants ［J］.Crop Science, 2005, 45(2): 437-448
［17］Zhan H X, Hodson J N, Williams J P, et al.Engineering salt-tolerant Brassica plants: characterization of yield and seed oil quality in transgenic plants with increased vacuolar sodium accumulation ［J］. PNAS Proceedings of the National Academy of Sciences, 2001, 98(22): 12832-12836
［18］Wu Y Y, Chen Q J, Chen M.Salt-tolerant transgenic perennial ryegrass (Lolium perenne L) obtained by Agrobacterium tumefaciens mediated transformation of the vacuolar Na+/H+?antiporter gene ［J］ Plant Science, 2005, 169: 65-73
［19］Mittler R. Oxidative stress, antioxidants and stress tolerance ［J］. Trends Plant Science, 2002, 7(9): 405-410
［20］Xiong L M, Schumaker K S, Zhu J K.Cell signaling during cold, drought, and salt stress ［J］.Plant Cell, 2002,14: 165-83.

备注/Memo

基金项目：河北大学引进人才启动项目(2011-221)；保定市科学技术研究与发展指导计划项目（11ZF096）；生物学强势特色学科资助。

第一作者简介：董丽君（1981-），女，硕士，主要从事分子遗传学研究。E-mail:donglijun1024@163.com。
通讯作者：刘建凤（1979-），女，博士，副教授，主要从事植物分子育种研究。E-mail：jianfengliu@hbu.edu.cn。

更新日期/Last Update: 2015-07-15