دورية أكاديمية
Pairwise growth competition assay for determining the replication fitness of human immunodeficiency viruses.
| العنوان: | Pairwise growth competition assay for determining the replication fitness of human immunodeficiency viruses. |
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| المؤلفون: | Manocheewa S; Department of Microbiology, University of Washington., Lanxon-Cookson EC; Department of Microbiology, University of Washington., Liu Y; Department of Microbiology, University of Washington., Swain JV; Department of Microbiology, University of Washington., McClure J; Department of Microbiology, University of Washington., Rao U; Department of Microbiology, University of Washington., Maust B; Department of Microbiology, University of Washington., Deng W; Department of Microbiology, University of Washington., Sunshine JE; Department of Microbiology, University of Washington., Kim M; Department of Microbiology, University of Washington., Rolland M; U.S Military HIV Research Program, Walter Reed Army Institute of Research; Henry M. Jackson Foundation., Mullins JI; Department of Microbiology, University of Washington; Departments of Medicine and Laboratory Medicine, University of Washington; jmullins@u.washington.edu. |
| المصدر: | Journal of visualized experiments : JoVE [J Vis Exp] 2015 May 04 (99), pp. e52610. Date of Electronic Publication: 2015 May 04. |
| نوع المنشور: | Journal Article; Research Support, N.I.H., Extramural; Video-Audio Media |
| اللغة: | English |
| بيانات الدورية: | Publisher: MYJoVE Corporation Country of Publication: United States NLM ID: 101313252 Publication Model: Electronic Cited Medium: Internet ISSN: 1940-087X (Electronic) Linking ISSN: 1940087X NLM ISO Abbreviation: J Vis Exp Subsets: MEDLINE |
| أسماء مطبوعة: | Original Publication: [Boston, Mass. : MYJoVE Corporation, 2006]- |
| مواضيع طبية MeSH: | HIV-1/*physiology , Virus Replication/*physiology, DNA Replication/genetics ; DNA Replication/physiology ; DNA, Viral/biosynthesis ; DNA, Viral/genetics ; HEK293 Cells ; HIV Core Protein p24/genetics ; HIV-1/genetics ; Humans ; Mutation ; Virus Replication/genetics |
| مستخلص: | In vitro fitness assays are essential tools for determining viral replication fitness for viruses such as HIV-1. Various measurements have been used to extrapolate viral replication fitness, ranging from the number of viral particles per infectious unit, growth rate in cell culture, and relative fitness derived from multiple-cycle growth competition assays. Growth competition assays provide a particularly sensitive measurement of fitness since the viruses are competing for cellular targets under identical growth conditions. There are several experimental factors to consider when conducting growth competition assays, including the multiplicity of infection (MOI), sampling times, and viral detection and fitness calculation methods. Each factor can affect the end result and hence must be considered carefully during the experimental design. The protocol presented here includes steps from constructing a new recombinant HIV-1 clone to performing growth competition assays and analyzing the experimental results. This protocol utilizes experimental parameter values previously shown to yield consistent and robust results. Alternatives are discussed, as some parameters need to be adjusted according to the cell type and viruses being studied. The protocol contains two alternative viral detection methods to provide flexibility as the availability of instruments, reagents and expertise varies between laboratories. |
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| معلومات مُعتمدة: | R01AI047734 United States AI NIAID NIH HHS; R01 AI047734 United States AI NIAID NIH HHS; R37 AI047734 United States AI NIAID NIH HHS; P01 AI057005 United States AI NIAID NIH HHS; R01AI111806 United States AI NIAID NIH HHS; P30 AI027757 United States AI NIAID NIH HHS; R01 AI111806 United States AI NIAID NIH HHS; P01AI057005 United States AI NIAID NIH HHS; R01 AI125026 United States AI NIAID NIH HHS |
| المشرفين على المادة: | 0 (DNA, Viral) 0 (HIV Core Protein p24) |
| تواريخ الأحداث: | Date Created: 20150521 Date Completed: 20151215 Latest Revision: 20190610 |
| رمز التحديث: | 20221213 |
| مُعرف محوري في PubMed: | PMC4542137 |
| DOI: | 10.3791/52610 |
| PMID: | 25993602 |
| قاعدة البيانات: | MEDLINE |
| تدمد: | 1940-087X |
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| DOI: | 10.3791/52610 |