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[Construction and expression of anti-clenbuterol single chain Fv recombinant vector].
Sheng Wu Gong Cheng Xue Bao. 2008 Aug; 24(8):1470-4.SW

Abstract

To construct the recombinant vector pBV220-scFv and express anti-clenbuterol (CBL) scFv antibody in Escherichia coli, we amplified the scFv gene using plasmid pCANTABSE-CBL as a template, recombined it with pPICZalphaA, then amplified the scFv-His-tag gene from plasmid pPICZalphaA-scFv and linked it with expression plasmid pBV220. We identified the recombinant plasmid by restrictive enzyme digestion, PCR amplification and sequence analysis. Finally, we transformed the recombinant vector into E. coli DH5alpha that was temperature-induced and expressed recombinant protein. We identified the recombinant protein by SDS-PAGE, Western blotting and indirect competitive ELISA. The results show that recombinant plasmid pBV220-scFv contained the inserted fragment with highest homology about 99.8%. The expression of scFv induced by temperature show 37 kD Mw and anti-His-tag mAb recognized-activity by SDS-PAGE and Western blotting respectively, and could competitively combine with CBL, the IC50 is 4.55 ng/mL. The recombinant plasmid pBV220-scFv is constructed and expresses the scFv gene of CBL in E. coli successfully. This study suggests the corresponding immunoassay methods could be set up by the recombinant scFv.

Authors+Show Affiliations

The Higher Education Key Laboratory of Food Quality and Safety of Guangdong Province/Research Institute of Food Quality and Safety of SCAU, South China Agricultural University, Guangzhou 510640, China. gzwhongd@163.comNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

English Abstract
Journal Article
Research Support, Non-U.S. Gov't

Language

chi

PubMed ID

18998553

Citation

Wang, Hong, et al. "[Construction and Expression of Anti-clenbuterol Single Chain Fv Recombinant Vector]." Sheng Wu Gong Cheng Xue Bao = Chinese Journal of Biotechnology, vol. 24, no. 8, 2008, pp. 1470-4.
Wang H, Liang Y, Yang J, et al. [Construction and expression of anti-clenbuterol single chain Fv recombinant vector]. Sheng Wu Gong Cheng Xue Bao. 2008;24(8):1470-4.
Wang, H., Liang, Y., Yang, J., Liu, X., Zhang, H., Lei, H., Shen, Y., & Sun, Y. (2008). [Construction and expression of anti-clenbuterol single chain Fv recombinant vector]. Sheng Wu Gong Cheng Xue Bao = Chinese Journal of Biotechnology, 24(8), 1470-4.
Wang H, et al. [Construction and Expression of Anti-clenbuterol Single Chain Fv Recombinant Vector]. Sheng Wu Gong Cheng Xue Bao. 2008;24(8):1470-4. PubMed PMID: 18998553.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - [Construction and expression of anti-clenbuterol single chain Fv recombinant vector]. AU - Wang,Hong, AU - Liang,Yan, AU - Yang,Jingyi, AU - Liu,Xixia, AU - Zhang,Hongbin, AU - Lei,Hongtao, AU - Shen,Yudong, AU - Sun,Yuanming, PY - 2008/11/13/pubmed PY - 2009/9/25/medline PY - 2008/11/13/entrez SP - 1470 EP - 4 JF - Sheng wu gong cheng xue bao = Chinese journal of biotechnology JO - Sheng Wu Gong Cheng Xue Bao VL - 24 IS - 8 N2 - To construct the recombinant vector pBV220-scFv and express anti-clenbuterol (CBL) scFv antibody in Escherichia coli, we amplified the scFv gene using plasmid pCANTABSE-CBL as a template, recombined it with pPICZalphaA, then amplified the scFv-His-tag gene from plasmid pPICZalphaA-scFv and linked it with expression plasmid pBV220. We identified the recombinant plasmid by restrictive enzyme digestion, PCR amplification and sequence analysis. Finally, we transformed the recombinant vector into E. coli DH5alpha that was temperature-induced and expressed recombinant protein. We identified the recombinant protein by SDS-PAGE, Western blotting and indirect competitive ELISA. The results show that recombinant plasmid pBV220-scFv contained the inserted fragment with highest homology about 99.8%. The expression of scFv induced by temperature show 37 kD Mw and anti-His-tag mAb recognized-activity by SDS-PAGE and Western blotting respectively, and could competitively combine with CBL, the IC50 is 4.55 ng/mL. The recombinant plasmid pBV220-scFv is constructed and expresses the scFv gene of CBL in E. coli successfully. This study suggests the corresponding immunoassay methods could be set up by the recombinant scFv. SN - 1000-3061 UR - https://www.unboundmedicine.com/medline/citation/18998553/[Construction_and_expression_of_anti_clenbuterol_single_chain_Fv_recombinant_vector]_ DB - PRIME DP - Unbound Medicine ER -