Chinese Journal of Tissue Engineering Research ›› 2011, Vol. 15 ›› Issue (33): 6177-6181.doi: 10.3969/j.issn.1673-8225.2011.33.024

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Construction and identification of lentiviral vector for Beclin1 gene

Wang Wen-yu1, 2, Fan Hong-kun3, Zhao Guo-qiang4, Qiao Peng3, Wang Shu-chun3, Wu Gang1   

  1. 1Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan  430023, Hubei Province, China；2Department of Medical Oncology, Henan Province People’s Hospital, Zhengzhou  450003, Henan Province, China; 3Department of Physiology, 4Department of Microbiology and Immunology, School of Medicine, Zhengzhou University, Zhengzhou 450052, Henan Province, China
  • Received:2011-03-08 Revised:2011-06-12 Online:2011-08-13 Published:2011-08-13
  • Contact: Wu Gang, Professor, Chief physician, Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430023, Hubei Province, China
  • About author:Wang Wen-yu☆, Studying for doctorate, Associate chief physician, Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430023, Hubei Province, China; Department of Medical Oncology, Henan Province People’s Hospital, Zhengzhou 450003, Henan Province, China wangwy0712@163.com
  • Supported by:

    the Science and Technology Innovation Talent Engineering Program of Henan Province, No.0623060900*

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Abstract:

BACKGROUND: Beclinl is an essential autophagy gene.
OBJECTIVE: To construct lentiviral vector pLenex-Beclin1 gene.
METHODS: Beclin1 gene amplification was used by real-time polymerase chain reaction. Gene amplification products were inserted in the lentiviral vector pLenex, and constructed lentiviral vector pLenex-Beclin1. Polymerase chain reaction analysis, double digests and DNA sequencing were used to confirm the constructed vectors whether or not success recombinant vector plasmid and to coinfect 293T cells, and then was transfected into non small cell lung cancer A549 cells. Western blot was then used to investigate the interfering efficiency of Beclin1 gene. 
RESULTS AND CONCLUSION: Polymerase chain reaction tests showed amplified positive fragments were inserted in pLenex vectors. Results of polymerase chain reaction analysis, double digestion and DNA sequencing showed that recombinant lentivirus plasmids pLenex-Beclin1 were constructed successfully. In transfected A549 cells, Beclin1 protein was over-expressed. Results verified that lentiviral vectors of Beclin1 gene over-expression were successfully constructed.

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