Journal of Integrative Agriculture ›› 2015, Vol. 14 ›› Issue (3): 430-437.DOI: 10.1016/S2095-3119(14)60919-9

• 论文 • 上一篇    下一篇

Enhanced resistance to Botrytis cinerea and Rhizoctonia solani in transgenic broccoli with a Trichoderma viride endochitinase gene

 YU Ya, ZHANG Lei, LIAN Wei-ran, XU Feng-feng, LI Shuang-tao, XIANG Juan, ZHANG Guo-zhen, HU Zan-min, ZHAO Bing, REN Shu-xin, GUO Yang-dong   

  1. 1、College of Agriculture & Biotechnology, China Agricultural University, Beijing 100193, P.R.China
    2、Department of Biology Science & Technology, Wuhan Institute of Bioengineering, Wuhan 430415, P.R.China
    3、Institute of Genetics & Developmental Biology, Chinese Academy of Sciences, Beijing 100101, P.R.China
    4、School of Agriculture, Virginia State University, Petersburg VA23806, USA
  • 收稿日期:2014-08-29 出版日期:2015-03-01 发布日期:2015-03-12
  • 通讯作者: GUO Yang-dong, Tel: +86-10-62734845,E-mail: yaguo@cau.edu.cn; ZHAO Bing, Tel: +86-10-62734095,E-mail: zhaobing@cau.edu.cn
  • 作者简介:* These authors contributed equally to this study.
  • 基金资助:

    This work was partly supported by the 863 Program (2012AA100A03), 973 Program (2012CB113900) and Beijing Agricultural Industry Project, China (BLVT-03).

Enhanced resistance to Botrytis cinerea and Rhizoctonia solani in transgenic broccoli with a Trichoderma viride endochitinase gene

 YU Ya, ZHANG Lei, LIAN Wei-ran, XU Feng-feng, LI Shuang-tao, XIANG Juan, ZHANG Guo-zhen, HU Zan-min, ZHAO Bing, REN Shu-xin, GUO Yang-dong     

  1. 1、College of Agriculture & Biotechnology, China Agricultural University, Beijing 100193, P.R.China
    2、Department of Biology Science & Technology, Wuhan Institute of Bioengineering, Wuhan 430415, P.R.China
    3、Institute of Genetics & Developmental Biology, Chinese Academy of Sciences, Beijing 100101, P.R.China
    4、School of Agriculture, Virginia State University, Petersburg VA23806, USA
  • Received:2014-08-29 Online:2015-03-01 Published:2015-03-12
  • Contact: GUO Yang-dong, Tel: +86-10-62734845,E-mail: yaguo@cau.edu.cn; ZHAO Bing, Tel: +86-10-62734095,E-mail: zhaobing@cau.edu.cn
  • About author:* These authors contributed equally to this study.
  • Supported by:

    This work was partly supported by the 863 Program (2012AA100A03), 973 Program (2012CB113900) and Beijing Agricultural Industry Project, China (BLVT-03).

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摘要： A endochitinase gene (Tch) from the fungus Trichoderma viride was introduced into broccoli (Brassica oleracea var. italica) by Agrobacterium-mediated transformation. Sixty-eight putative transformants were obtained and the presence of the Tch gene was confirmed by both PCR and Southern blot analysis. RT-PCR analysis showed an accumulation of the transcript encoding the endochitinase protein in the transgenic plants. Using real-time quantitative PCR, the expression profiling of endochitinase gene was analyzed. Primary transformants and selfed progeny were examined for expression of the endochitinase using a fluorometric assay and for their resistance to the pathogenic fungi Botrytis cinerea and Rhizoctonia solani. The endochitinase activities in T0 in vitro plants, T0 mature plants and T1 mature plants were correlated with leaf lesions, and the transgenic line T618 had high endochitinse activities of 102.68, 114.53 and 120.27 nmol L–1 MU min–1 mg–1 protein in the three kinds of plants, respectively. The endochitinase activity showed a positive correlation with the resistance to the pathogens. Most transgenic T0 broccoli had increased resistance to the pathogens of B. cinerea and R. solani in leaf assays and this resistance was confirmed to be inheritable. These findings suggested that expression of the Tch gene from T. viride could enhance resistance to pathogenic fungi in Brassica species.

关键词: Botrytis cinerea , broccoli , endochitinase gene , genetic transformation , Rhizoctonia solani

Abstract: A endochitinase gene (Tch) from the fungus Trichoderma viride was introduced into broccoli (Brassica oleracea var. italica) by Agrobacterium-mediated transformation. Sixty-eight putative transformants were obtained and the presence of the Tch gene was confirmed by both PCR and Southern blot analysis. RT-PCR analysis showed an accumulation of the transcript encoding the endochitinase protein in the transgenic plants. Using real-time quantitative PCR, the expression profiling of endochitinase gene was analyzed. Primary transformants and selfed progeny were examined for expression of the endochitinase using a fluorometric assay and for their resistance to the pathogenic fungi Botrytis cinerea and Rhizoctonia solani. The endochitinase activities in T0 in vitro plants, T0 mature plants and T1 mature plants were correlated with leaf lesions, and the transgenic line T618 had high endochitinse activities of 102.68, 114.53 and 120.27 nmol L–1 MU min–1 mg–1 protein in the three kinds of plants, respectively. The endochitinase activity showed a positive correlation with the resistance to the pathogens. Most transgenic T0 broccoli had increased resistance to the pathogens of B. cinerea and R. solani in leaf assays and this resistance was confirmed to be inheritable. These findings suggested that expression of the Tch gene from T. viride could enhance resistance to pathogenic fungi in Brassica species.

Key words: Botrytis cinerea , broccoli , endochitinase gene , genetic transformation , Rhizoctonia solani