title: Cell Compatibility of an Eposimal Vector Mediated by the Characteristic Motifs of Matrix Attachment Regions.
abstract: The characteristic sequence of β-interferon matrix attachment regions (MARs) can mediate transgene expression via episomal vectors in Chinese hamster ovary (CHO) cells. However, the host cells were from hamster ovaries, which are not suitable target cells for gene therapy. In this study, we aimed to evaluate the suitability of 12 different human cell lines as target cells for gene therapy. We transfected the cells with episomal vectors and obtained colonies stably expressing the vector products after G418 screening. Therefore the stably transfected cells were split into two and further cultured either in the presence or the absence of G418. Flow cytometry was used to observe the positive rate of cell transfection and level of green fluorescent protein (GFP) expression. Plasmid rescue assays, fluorescence in situ hybridization (FISH), and fluorescence quantitative polymerase chain reaction (PCR) were used to investigate the presence and gene copy numbers of plasmid in mammalian cells. The results showed that transfection efficiency and transgene expression levels in A375, Eca-109, and Changliver cells were high. In contrast, transgene silencing was observed in BJ, HSF, and A431 cells, and low expression of the transgene was observed in the other six cell lines. In addition, the plasmid was present in the episomal state in A375, Eca-109, and Chang-liver cells with relatively low copy numbers even under nonselective conditions. Thus, our results provide the first evidence showing transgene expression of an episomal vector mediated by the characteristic motifs of MARs for maintenance of the longterm stability of episomes in different types of cells.
authors.forename: Tian-Yun
authors.initials: TY
authors.lastname: Wang
authors.forename: Li
authors.initials: L
authors.lastname: Wang
authors.forename: Yu-Xin
authors.initials: YX
authors.lastname: Yang
authors.forename: Chun-Peng
authors.initials: CP
authors.lastname: Zhao
authors.forename: Yan-Long
authors.initials: YL
authors.lastname: Jia
authors.forename: Qin
authors.initials: Q
authors.lastname: Li
authors.forename: Jun-He
authors.initials: JH
authors.lastname: Zhang
authors.forename: Yi-You
authors.initials: YY
authors.lastname: Peng
authors.forename: Miao
authors.initials: M
authors.lastname: Wang
authors.forename: Hong-Yan
authors.initials: HY
authors.lastname: Xu
authors.affiliation: Department of Biochemistry and Molecular Biology, Xinxiang Medical University, Jinsui Road, Xinxiang 453003, Henan, China.
authors.forename: Xiao-Yin
authors.initials: XY
authors.lastname: Wang
mesh_terms: D000818:Animals; D002460:Cell Line; D018628:Gene Dosage; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D045170:Matrix Attachment Regions / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods
chemical_list: D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins
keywords: Eposimal vector; Host cell; Matrix attachment regions; Transgene expression