title: Characterization of retroviral infectivity and superinfection resistance during retrovirus-mediated transduction of mammalian cells.
abstract: Retroviral vectors including lentiviral vectors are commonly used tools to stably express transgenes or RNA molecules in mammalian cells. Their utilities are roughly divided into two categories, stable overexpression of transgenes and RNA molecules, which requires maximal transduction efficiency, or functional selection with retrovirus (RV)-based libraries, which takes advantage of retroviral superinfection resistance. However, the dynamic features of RV-mediated transduction are not well characterized. Here, we engineered two murine stem cell virus-based retroviral vectors expressing dual fluorescence proteins and antibiotic markers, and analyzed virion production efficiency and virion stability, dynamic infectivity and superinfection resistance in different cell types, and strategies to improve transduction efficiency. We found that the highest virion production occurred between 60 and 72 h after transfection. The stability of the collected virion supernatant decreased by >60% after 3 days in storage. We found that RV infectivity varied drastically in the tested human cancer lines, while low transduction efficiency was partially overcome with increased virus titer, prolonged infection duration and/or repeated infections. Furthermore, we demonstrated that RV receptors PIT1 and PIT2 were lowly expressed in the analyzed cells, and that PIT1 and/or PIT2 overexpression significantly improved transduction efficiency in certain cell lines. Thus, our findings provide resourceful information for the optimal conditions of retroviral-mediated gene delivery.
authors.affiliation: Department of Orthopaedic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.|Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: J
authors.initials: J
authors.lastname: Liao
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Q
authors.initials: Q
authors.lastname: Wei
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.
authors.forename: J
authors.initials: J
authors.lastname: Fan
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Zou
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Conservative Dentistry and Endodontics, West China School of Stomatology, Sichuan University, Chengdu, China.
authors.forename: D
authors.initials: D
authors.lastname: Song
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Orthopaedic Surgery, Union Hospital of Tongji Medical College, Huazhong University of Science &Technology, Wuhan, China.
authors.forename: J
authors.initials: J
authors.lastname: Liu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: F
authors.initials: F
authors.lastname: Liu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Neurosurgery, The Affiliated Zhongnan Hospital of Wuhan University, Wuhan, China.
authors.forename: C
authors.initials: C
authors.lastname: Ma
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.|Department of Blood Transfusion, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China.
authors.forename: X
authors.initials: X
authors.lastname: Hu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Biomedical Engineering, School of Bioengineering, Chongqing University, Chongqing, China.
authors.forename: L
authors.initials: L
authors.lastname: Li
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Emergency Medicine, Beijing Hospital affiliated with Peking University Health Sciences Center, Beijing, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: X
authors.initials: X
authors.lastname: Qu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: L
authors.initials: L
authors.lastname: Chen
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: X
authors.initials: X
authors.lastname: Yu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Orthopaedic Surgery, Union Hospital of Tongji Medical College, Huazhong University of Science &Technology, Wuhan, China.
authors.forename: Z
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Department of Orthopaedic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.|Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: C
authors.initials: C
authors.lastname: Zhao
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.
authors.forename: Z
authors.initials: Z
authors.lastname: Zeng
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.
authors.forename: R
authors.initials: R
authors.lastname: Zhang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.
authors.forename: S
authors.initials: S
authors.lastname: Yan
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of ENT, The Affiliated Zhongnan Hospital of Wuhan University, Wuhan, China.
authors.forename: T
authors.initials: T
authors.lastname: Wu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: X
authors.initials: X
authors.lastname: Wu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Shu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: J
authors.initials: J
authors.lastname: Lei
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Li
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Department of Laboratory Medicine and Clinical Diagnostics, The Affiliated Yantai Hospital, Binzhou Medical University, Yantai, China.
authors.forename: W
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: J
authors.initials: J
authors.lastname: Wang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Section of Plastic Surgery, Department of Surgery, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: R R
authors.initials: RR
authors.lastname: Reid
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: M J
authors.initials: MJ
authors.lastname: Lee
authors.affiliation: Department of Orthopaedic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.
authors.forename: W
authors.initials: W
authors.lastname: Huang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Wolf
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.|Chinese Ministry of Education Key Laboratory of Diagnostic Medicine, Chongqing Medical University, Chongqing, China.|The Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: T-C
authors.initials: TC
authors.lastname: He
authors.affiliation: Department of Blood Transfusion, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China.
authors.forename: J
authors.initials: J
authors.lastname: Wang
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D011991:Receptors, Virus / Q000235:genetics / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D014162:Transfection / Q000379:methods* / Q000592:standards; D014771:Virion / Q000235:genetics / Q000378:metabolism
chemical_list: D011991:Receptors, Virus; D049452:Green Fluorescent Proteins