AUTHOR=Li Yiquan , Zhu Yilong , Chen Shuang , Li Wenjie , Yin Xunzhe , Li Shanzhi , Xiao Pengpeng , Han Jicheng , Li Xiao , Sun Lili , Jin Ningyi 

TITLE=Generation of an Attenuated Tiantan Vaccinia Virus Strain by Deletion of Multiple Genes

JOURNAL=Frontiers in Cellular and Infection Microbiology

VOLUME=7

YEAR=2017

URL=https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2017.00462

DOI=10.3389/fcimb.2017.00462

ISSN=2235-2988

ABSTRACT=<p>An attenuated vaccinia virus-MVTT<sub>EAB</sub>-was constructed by deletion of non-essential gene segments related to the immunomodulatory and virulence functions of the vaccinia virus Tiantan strain (VVTT). The shuttle plasmids pTC-EGFP, pTE-EGFP, pTA35-EGFP, pTB-EGFP, and pTA66-EGFP were constructed and combined with the early and late strong promoter pE/L and EGFP as an exogenous selectable marker. Then, through the homologous recombination technology and Cre/loxP system, the following gene fragments were gradually knocked out one by one: TC7L-TK2L, TE3L, TA35R, TB13R, and TA66R. Ultimately, the five-segment-deleted attenuated strain MVTT<sub>EAB</sub> was obtained. Knockout of these segments and genetic stability of MVTT<sub>EAB</sub> were confirmed, and it was also shown that knockout of these segments did not affect the replication ability of the virus. Further, a series of <italic>in vivo</italic> and <italic>in vitro</italic> experiments demonstrated that the virulence of MVTT<sub>EAB</sub> was attenuated significantly, but at same time, high immunogenicity was maintained. These results indicate that MVTT<sub>EAB</sub> has potential for clinical use as a safe viral vector or vaccine with good attenuation and immunogenicity.</p>