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| Construction of Recombinant Lentiviral Vector of shRNA for CHD5 Gene and the Study of Its Functions in Human Colon Carcinoma Lovo Cells |
| ZHAO Rui, LV Jing-ye, HUANG Hai-li, YAN Qi-tao |
| Institute of Genetic Engineering of Southern Medical University, Guangzhou 510515, China |
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Abstract To study the effects of lentiviral vector of microRNA targeting CHD5 gene on human colon carcinoma Lovo cell. Methods: Specific small interference RNAs were designed on line software and screened with the optimize principles, synthesized and cloned into the pPRIME vetor. Recombinant lentiviral was producted by 293FT cells following co-transfection of pPRIME-TET-GFP-CHD5 with the packaging plasmids pCMV-VSV-G, pRSV-Rev and pMDLg-pRRE. Human colon carcinoma Lovo cell were infected by the recombinant lentivirus.The expression of CHD5 was confirmed by RT-PCR and Western blotting, and the proliferation of Lovo cells was evaluated by MTT assay. Results: The recombinant lentiviral vecor carried the CHD5 gene was successfully constructed by restriction enzyme digestion and sequencing, respectively.The packaged lentiviral titer was 3.1×106 TU/ml.RT-PCR and Western blot analysis revealed that pPRIME-TET-GFP-CHD5 infection inhibited CHD5 mRNA and protein expressions in Lovo cells. The proliferation of Lovo cells was increased. Conclusion: The recombinant lentiviral vector pPRIME-TET-GFP-CHD5 was constructed successfully. It can be deliver target gene to human colon carcinoma Lovo cell, and inhibit CHD5 can promoted the proliferation of colon carcinoma Lovo cell.
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Received: 08 February 2012
Published: 25 May 2012
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