title: A method mediated AAVS1 recombination with Rep mRNA and homologous arms.
abstract: The adeno-associated virus (AAV) genome can be stably integrated into the AAVS1 region of human chromosome 19 (19q13.4-qter) with the assistance of Rep68/78 protein. In the current models of AAV integration in a locus-specific manner, the foreign genes were randomly inserted into the AAVS1 region, which contains several functional genes. As random integration in this region may lead to insertion mutations and disrupt normal gene expression or critical signaling pathways of the host cells, it is necessary to find a precise insertion site in the AAVS1 region. Homologous recombination is the most accurate and versatile mechanism for such site-specific integration. To investigate site-specific integration in the AAVS1 region, a targeted vector containing two homologous arms derived from AAVS1 and a reporter gene was transfected into HeLa cells with or without Rep68/78 mRNA. The results indicated that transient expression of Rep68/78 in HeLa cells improved integration of the gene of interest at the AAVS1 locus in a site-specific manner. Compared with locus-specific integration reported in previous studies, site-specific integration may minimize the risk associated with random DNA integration in the AAVS1 region, which might be helpful for gene therapy.
authors.affiliation: Department of Biochemistry and Molecular Biology,  Anhui Medical University, Hefei 230032,  China.
authors.forename: Qiantong
authors.initials: Q
authors.lastname: Xiang
authors.forename: Linian
authors.initials: L
authors.lastname: Huang
authors.forename: Sijia
authors.initials: S
authors.lastname: Guo
authors.forename: Fang
authors.initials: F
authors.lastname: Chen
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Zha
authors.forename: Bing
authors.initials: B
authors.lastname: Chen
authors.forename: Liangdan
authors.initials: L
authors.lastname: Sun
authors.forename: Haisheng
authors.initials: H
authors.lastname: Zhou
authors.forename: Depei
authors.initials: D
authors.lastname: Liu
mesh_terms: D001665:Binding Sites / Q000235:genetics; D002888:Chromosomes, Human, Pair 19 / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D005434:Flow Cytometry; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008957:Models, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D014161:Transduction, Genetic / Q000379:methods; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D016662:Virus Integration
chemical_list: D004268:DNA-Binding Proteins; D012333:RNA, Messenger; D014764:Viral Proteins; C068734:rep proteins, Adeno-associated virus 2; D049452:Green Fluorescent Proteins