Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

doi:10.1016/S1671-2927(00)8543

Journal of Integrative Agriculture ›› 2012, Vol. 11 ›› Issue (2): 263-268.DOI: 10.1016/S1671-2927(00)8543

Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

WU Jian-xiang, SHANG Hai-li, XIE Yan, SHEN Qing-tang , ZHOU Xue-ping

1.Institute of Biotechnology, Zhejiang University, Hangzhou 310029, P.R.China

收稿日期:2011-03-01 出版日期:2012-02-01 发布日期:2012-02-11
通讯作者: Correspondence ZHOU Xue-ping, Tel: +86-571-88982680, Fax: +86-571-86971498, E-mail: zzhou@zju.edu.cn
基金资助:
This work was supported by the Special Fund for Agro- Scientific Research in the Public Interest from the Ministry of Agriculture, China (201003065).

Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

WU Jian-xiang, SHANG Hai-li, XIE Yan, SHEN Qing-tang , ZHOU Xue-ping

1.Institute of Biotechnology, Zhejiang University, Hangzhou 310029, P.R.China

Received:2011-03-01 Online:2012-02-01 Published:2012-02-11
Contact: Correspondence ZHOU Xue-ping, Tel: +86-571-88982680, Fax: +86-571-86971498, E-mail: zzhou@zju.edu.cn
Supported by:
This work was supported by the Special Fund for Agro- Scientific Research in the Public Interest from the Ministry of Agriculture, China (201003065).

摘要/Abstract

摘要： Tomato yellow leaf curl virus (TYLCV) is a species of the family Geminiviridae, causing serious yield losses in tomato production. The coat protein (CP) gene of TYLCV isolate SH2 was expressed in Escherichia coli BL21 (DE3) using pET- 32a as the expression vector. The recombinant protein was purified through Ni+-NTA affinity column and used to immunize BALB/c mice. Three hybridoma cell lines (2B2, 2E3 and 3E10) secreting monoclonal antibodies (MAbs) against TYLCV CP were obtained by fusing mouse myeloma cells (Sp 2/0) with spleen cells from the immunized BALB/c mouse. The titers of ascitic fluids of three MAbs ranged from 10-6 to 10-7 in indirect-ELISA. Isotypes and subclasses of all the MAbs belonged to IgG1, κ light chain. Triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) showed that the MAb 3E10 could react with five begomoviruses infecting tomato, while the other two (2B2 and 2E3) mainly reacted with TYLCV. TAS-ELISA was set up using the MAb 3E10, and the established method could successfully detect virus in plant sap at 1:2 560 (w/v, g mL-1). Detection of field samples showed that begomoviruses were common in tomato crops in Zhejiang Province, China.

关键词: Tomato yellow leaf curl virus, coat protein, monoclonal antibody, TAS-ELISA

Abstract: Tomato yellow leaf curl virus (TYLCV) is a species of the family Geminiviridae, causing serious yield losses in tomato production. The coat protein (CP) gene of TYLCV isolate SH2 was expressed in Escherichia coli BL21 (DE3) using pET- 32a as the expression vector. The recombinant protein was purified through Ni+-NTA affinity column and used to immunize BALB/c mice. Three hybridoma cell lines (2B2, 2E3 and 3E10) secreting monoclonal antibodies (MAbs) against TYLCV CP were obtained by fusing mouse myeloma cells (Sp 2/0) with spleen cells from the immunized BALB/c mouse. The titers of ascitic fluids of three MAbs ranged from 10-6 to 10-7 in indirect-ELISA. Isotypes and subclasses of all the MAbs belonged to IgG1, κ light chain. Triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) showed that the MAb 3E10 could react with five begomoviruses infecting tomato, while the other two (2B2 and 2E3) mainly reacted with TYLCV. TAS-ELISA was set up using the MAb 3E10, and the established method could successfully detect virus in plant sap at 1:2 560 (w/v, g mL-1). Detection of field samples showed that begomoviruses were common in tomato crops in Zhejiang Province, China.

Key words: Tomato yellow leaf curl virus, coat protein, monoclonal antibody, TAS-ELISA

WU Jian-xiang, SHANG Hai-li, XIE Yan, SHEN Qing-tang , ZHOU Xue-ping. Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection[J]. Journal of Integrative Agriculture, 2012, 11(2): 263-268.

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Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

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