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中国生物工程杂志

China Biotechnology
China Biotechnology  2013, Vol. 33 Issue (10): 36-43    DOI:
    
Expression of Herpes Simplex Virus Type 2 Latency-associated Transcript Open Reading Frames and Its Anti-apoptotic Effects
ZHONG Fei-fei1,2, YANG Hui-lan1, LÜ Fang-biao1,2, XUE Li-zhang1,2, BAI Li-li1,2, FAN Jian-yong1
1. Department of Dermatology, Guangzhou Liuhuaqiao Hospital, Guangzhou 510010, China;
2. School of Biological Science and Engineering, South China University of Technology, Guangzhou 510006, China
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Abstract  Objective:To study the effects of herpes simplex virus type 2(HSV-2) latency-associated transcript(LAT) open reading frames(ORFs) on Vero cells which were induced apoptosis by agent. Methods:PCR amplified LAT ORF1, ORF2, ORF3 fragment,then construct recombinant plasmid pEGFP-ORF1,pEGFP-ORF2,pEGFP-ORF3,transfect them into Vero cells and identify its expression by RT-PCR.Using actinomycin D,5-FU and cisplatinere to induce Vero cells apoptosis,the changes of Vero cells morphology was observed by fluorescence microscopy.MTT assay was used to evaluate Vero cells viability and flow cytometry was used to calculate the cells apoptosis rate. Results:The pEGFP-ORF1,pEGFP-ORF2,pEGFP-ORF3 was constructed successfully which was confirmed by Double digestion and sequencing. Vero cells morphology were normal when transfected with pEGFP-ORFs and induced by apoptosis-inducing agent. MTT assay showed that the viabilities of Vero cells in recombinant plasmid group were no significant difference compared with the normal control group(P> 0.05), but remarkable higher than empty plasmid pEGFP-C2 group,the difference was significant(P<0.05).Flow cytometry assay showed that the cells apoptosis rate in pEGFP-ORFs group were no significant difference compared with the normal control group(P> 0.05),but was remarkable lower than the pEGFP-C2 group(P<0.05). Conclusion HSV-2 LAT ORFs gene can effectively expressed in Vero cells and can protect Vero cells from apoptosis.

Key wordsORFs      Herpes simplex virus      LAT      Apoptosis-inducing agent      Apoptosis      Vero cells     
Received: 19 July 2013      Published: 25 October 2013
ZTFLH:  Q819  
Cite this article:

ZHONG Fei-fei, YANG Hui-lan, LÜ Fang-biao, XUE Li-zhang, BAI Li-li, FAN Jian-yong. Expression of Herpes Simplex Virus Type 2 Latency-associated Transcript Open Reading Frames and Its Anti-apoptotic Effects. China Biotechnology, 2013, 33(10): 36-43.

URL:

http://manu60.magtech.com.cn/biotech/     OR     http://manu60.magtech.com.cn/biotech/Y2013/V33/I10/36

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