Protocol for high-throughput reservoir quantification across global HIV subtypes using a cross-subtype intact proviral DNA assay

Autor: Carolyn S. Fish, Noah A.J. Cassidy, Claire N. Levy, Sean M. Hughes, Keith R. Jerome, Julie Overbaugh, Florian Hladik, Dara A. Lehman
Jazyk: angličtina
Rok vydání: 2022
Předmět:
Zdroj: STAR Protocols, Vol 3, Iss 4, Pp 101681- (2022)
Druh dokumentu: article
ISSN: 2666-1667
DOI: 10.1016/j.xpro.2022.101681
Popis: Summary: The cross-subtype intact proviral DNA assay (CS-IPDA) is a high-throughput method to quantify HIV reservoir size in populations infected with any of the dominant global HIV-1 subtypes. Our protocol includes genomic DNA isolation optimized to minimize DNA shearing, a reference droplet digital PCR (ddPCR) assay to quantify T cells and assess DNA shearing, and a multiplex ddPCR targeting three distinct regions across the HIV genome to quantify intact proviruses as an estimate of replication-competent proviruses in the reservoir.For complete details on the use and execution of this protocol, please refer to Cassidy et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
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