转hrpZ_(psta)和chi双价广谱抗病基因大豆对疫霉根腐病的抗性分析
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摘要
疫霉根腐病能使大豆感病品种减产5%～10%,并造成品质降低。利用转基因技术培育抗病品种是解决该问题的有效途径之一。对转hrpZ_(psta)和chi双价广谱抗病基因株系T2003-23-11、T2004-28-321和T2004-30-384进行分子检测并鉴定其抗疫霉根腐病能力。Southern Blot结果表明,hrpZ_(psta)和chi双价基因已整合在株系的核基因组中而且稳定遗传到T_4代;qRT-PCR结果表明,hrpZ_(psta)和chi双价基因在株系的不同组织中的相对表达量均高于非转基因对照。利用下胚轴侵染法对3个株系进行抗疫霉根腐病鉴定结果表明,3个株系抗病级别均为高抗,非转基因对照的抗病级别为中抗,表明转基因株系的抗病能力得到提高。
Phytophthora root rot can make yield of sensitive soybean cultivar decrease by 5%～10% and quality decrease, so cultivating disease-resistant varieties by transgenic technology becomes the effective way of solving the problem. Molecular detection of strains T2003-23-11, T2004-28-321 and T2004-30-384 was conducted and their disease-resistant ability was identified with two broad spectrum disease-resistant genes hrpZ_(psta) and chi. Southern Blot results demonstrate hrpZpsta and chi genes have integrated in the nuclear genome and stably inherited to T_4 generation. qRT-PCR results demonstrate expression in different tissues of strains are higher than that of the non-transgenic control. Phytophthora root rot of three strains was identified by hypocotyl infection method,and the results show that resistance level of the three strains is high, and resistance level of non-transgenic control is moderate, showing that disease resistance of transgenic strains is improved.
Phytophthora root rot can make yield of sensitive soybean cultivar decrease by 5%～10% and quality decrease, so cultivating disease-resistant varieties by transgenic technology becomes the effective way of solving the problem. Molecular detection of strains T2003-23-11, T2004-28-321 and T2004-30-384 was conducted and their disease-resistant ability was identified with two broad spectrum disease-resistant genes hrpZ_(psta) and chi. Southern Blot results demonstrate hrpZpsta and chi genes have integrated in the nuclear genome and stably inherited to T_4 generation. qRT-PCR results demonstrate expression in different tissues of strains are higher than that of the non-transgenic control. Phytophthora root rot of three strains was identified by hypocotyl infection method,and the results show that resistance level of the three strains is high, and resistance level of non-transgenic control is moderate, showing that disease resistance of transgenic strains is improved.
引文
[1] 周恩远, 刘丽君, 祖伟,等. 春大豆农艺性状与品质相关系的研究[J]. 东北农业大学报, 2008, 39: 145-149.
[2] 刘清玮,高延辉,王希.大豆皂苷脂质体的制备及稳定性研究[J]. 吉林农业大学学报,2017,39(6):619-623.
[3] 刘子强,王有江. 浅谈大豆常见病害症状识别与防治[J]. 黑龙江科技信息,2010,21:240.
[4] 芮海英. 几丁质酶活性与大豆抗疫霉根腐病的关系[D].大庆:黑龙江八一农垦大学,2008.
[5] Lindgren P B.The role of hrp genes during plant-bacterial interactions[J].Annu Rev Phy Topathol,1997,35:129-152.
[6] 张云月,付永平,王丕武,等. 转hrpZpsta抗病基因大豆的研究[J]. 西北农林科技大学学报(自然科学版),2011,39(9):86-92.
[7] Wei Z M,Beer S V.Harpin from Erwinia amylovora induces plant resistance[J]. Acta Horticulture, 1996, 411:223-225.
[8] 尹俊琦. 转hrpZpsta基因大豆后代的遗传稳定性研究[D].长春:吉林农业大学,2013.
[9] 季丽丽. 玉米遗传转化体系的优化及HrpZ基因转化玉米的研究[D].长春:吉林农业大学,2011.
[10] 谷晓娜,刘振库,王丕武,等. hrpZ_(Psta)在转基因大豆中定量表达与疫霉根腐病和灰斑病抗性相关研究[J]. 中国油料作物学报,2015,37(1):35-40.
[11] 左豫虎,芮海英,杨传平,等. 几丁质酶活性与大豆抗疫霉根腐病的关系[J]. 植物保护,2008,34(6):49-53.
[12] 谢树章,刘亚娟,秦平伟,等.植物几丁质酶及应用研究进展[J].安徽农学通,2009,15(8):58-61.
[13] 胡春华,魏岳荣,刘凯,等. 几丁质酶基因克隆及其野生蕉转化[J]. 分子植物育种,2010,8(4):719-724.
[14] 王全伟,曲敏,张海玲,等. 菜豆几丁质酶基因Bchi的克隆及其在转基因烟草中的表达[J]. 分子植物育种,2008,6(1):53-58.
[15] 程红梅,简桂良,倪万潮,等. 转几丁质酶和β-1,3-葡聚糖酶基因提高棉花对枯萎病和黄萎病的抗性[J]. 中国农业科学,2005,38(6):1160-1166.
[16] 卢实,王丕武,曲静,等. 广谱抗病基因chi和hrpZpsta双价表达载体的构建及其在大豆中的遗传转化[J]. 大豆科学,2015,34(1):26-31.
[17] 郭玉双,张艳菊,朱延明,等. 抗真菌转基因大豆对大豆疫霉根腐病抗病性鉴定[J]. 东北农业大学学报,2006,37(6):733-738.
[18] 宋阳. NPR1和HrpZpsg12双价抗病基因植物表达载体的构建及在大豆中的遗传转化[D].长春:吉林农业大学,2014.
[19] 黄珏. 转几丁质酶基因和β-1,3-葡聚糖酶基因玉米抗纹枯病研究[D].武汉:华中农业大学,2013.
[20] 张昕,金中时,王凤良,等. 转几丁质酶和葡聚糖酶双价基因棉花株系的抗病性[J]. 江苏农业科学,2012,40(12):130-132.
[21] 姚丹,王丕武,刘占柱,等.农杆菌介导法将Bt杀虫蛋白基因导入玉米自交系的研究[J].吉林农业大学学报,2004,26(1):27-31.
[22] 关淑艳,王丕武,马义勇,等.应用农杆菌介导法将淀粉分支酶基因反义表达载体转入玉米自交系的研究[J].吉林农业大学学报,2005,27(5):498-502.
[23] 王春艳,关淑艳,王丕武,等.农杆菌介导的BADH-CMO基因转化玉米愈伤组织的初步研究[J].吉林农业大学学报,2012,34(5):503-507,510.
[24] 何欢,高嵩,韩丹,等.抗虫基因Cry1lem转化大豆的研究[J].吉林农业大学学报,2017,39(6):665-669.
[25] 魏洪波,王丕武,张卓,等.大豆查尔酮还原酶GmCHR基因的克隆及其在烟草中的表达[J].吉林农业大学学报,2014,36(5):546-553.
[26] 华志华, 朱雪峰,吴明国,等.水稻转基因整合模式中外源基因的遗传规律[J]. 作物学报, 2003,29(1): 44-48.
[27] Li X G, Chen S B, Lu Z X,et al. Impact of copynumber on transgenic expression intobacco[J].Atca Botanica Sinica,2002,44 (1):120-123.
[28] Bavage A D, Buck E, Dale P,et al.Analysis of a backcross population from a multi-copy transgenic Brassica napus lin[J]. Euphytica, 2002,124(3): 333-340.
[2] 刘清玮,高延辉,王希.大豆皂苷脂质体的制备及稳定性研究[J]. 吉林农业大学学报,2017,39(6):619-623.
[3] 刘子强,王有江. 浅谈大豆常见病害症状识别与防治[J]. 黑龙江科技信息,2010,21:240.
[4] 芮海英. 几丁质酶活性与大豆抗疫霉根腐病的关系[D].大庆:黑龙江八一农垦大学,2008.
[5] Lindgren P B.The role of hrp genes during plant-bacterial interactions[J].Annu Rev Phy Topathol,1997,35:129-152.
[6] 张云月,付永平,王丕武,等. 转hrpZpsta抗病基因大豆的研究[J]. 西北农林科技大学学报(自然科学版),2011,39(9):86-92.
[7] Wei Z M,Beer S V.Harpin from Erwinia amylovora induces plant resistance[J]. Acta Horticulture, 1996, 411:223-225.
[8] 尹俊琦. 转hrpZpsta基因大豆后代的遗传稳定性研究[D].长春:吉林农业大学,2013.
[9] 季丽丽. 玉米遗传转化体系的优化及HrpZ基因转化玉米的研究[D].长春:吉林农业大学,2011.
[10] 谷晓娜,刘振库,王丕武,等. hrpZ_(Psta)在转基因大豆中定量表达与疫霉根腐病和灰斑病抗性相关研究[J]. 中国油料作物学报,2015,37(1):35-40.
[11] 左豫虎,芮海英,杨传平,等. 几丁质酶活性与大豆抗疫霉根腐病的关系[J]. 植物保护,2008,34(6):49-53.
[12] 谢树章,刘亚娟,秦平伟,等.植物几丁质酶及应用研究进展[J].安徽农学通,2009,15(8):58-61.
[13] 胡春华,魏岳荣,刘凯,等. 几丁质酶基因克隆及其野生蕉转化[J]. 分子植物育种,2010,8(4):719-724.
[14] 王全伟,曲敏,张海玲,等. 菜豆几丁质酶基因Bchi的克隆及其在转基因烟草中的表达[J]. 分子植物育种,2008,6(1):53-58.
[15] 程红梅,简桂良,倪万潮,等. 转几丁质酶和β-1,3-葡聚糖酶基因提高棉花对枯萎病和黄萎病的抗性[J]. 中国农业科学,2005,38(6):1160-1166.
[16] 卢实,王丕武,曲静,等. 广谱抗病基因chi和hrpZpsta双价表达载体的构建及其在大豆中的遗传转化[J]. 大豆科学,2015,34(1):26-31.
[17] 郭玉双,张艳菊,朱延明,等. 抗真菌转基因大豆对大豆疫霉根腐病抗病性鉴定[J]. 东北农业大学学报,2006,37(6):733-738.
[18] 宋阳. NPR1和HrpZpsg12双价抗病基因植物表达载体的构建及在大豆中的遗传转化[D].长春:吉林农业大学,2014.
[19] 黄珏. 转几丁质酶基因和β-1,3-葡聚糖酶基因玉米抗纹枯病研究[D].武汉:华中农业大学,2013.
[20] 张昕,金中时,王凤良,等. 转几丁质酶和葡聚糖酶双价基因棉花株系的抗病性[J]. 江苏农业科学,2012,40(12):130-132.
[21] 姚丹,王丕武,刘占柱,等.农杆菌介导法将Bt杀虫蛋白基因导入玉米自交系的研究[J].吉林农业大学学报,2004,26(1):27-31.
[22] 关淑艳,王丕武,马义勇,等.应用农杆菌介导法将淀粉分支酶基因反义表达载体转入玉米自交系的研究[J].吉林农业大学学报,2005,27(5):498-502.
[23] 王春艳,关淑艳,王丕武,等.农杆菌介导的BADH-CMO基因转化玉米愈伤组织的初步研究[J].吉林农业大学学报,2012,34(5):503-507,510.
[24] 何欢,高嵩,韩丹,等.抗虫基因Cry1lem转化大豆的研究[J].吉林农业大学学报,2017,39(6):665-669.
[25] 魏洪波,王丕武,张卓,等.大豆查尔酮还原酶GmCHR基因的克隆及其在烟草中的表达[J].吉林农业大学学报,2014,36(5):546-553.
[26] 华志华, 朱雪峰,吴明国,等.水稻转基因整合模式中外源基因的遗传规律[J]. 作物学报, 2003,29(1): 44-48.
[27] Li X G, Chen S B, Lu Z X,et al. Impact of copynumber on transgenic expression intobacco[J].Atca Botanica Sinica,2002,44 (1):120-123.
[28] Bavage A D, Buck E, Dale P,et al.Analysis of a backcross population from a multi-copy transgenic Brassica napus lin[J]. Euphytica, 2002,124(3): 333-340.