Single Genome Sequencing of Expressed and Proviral HIV-1 Envelope Glycoprotein 120 (gp120) and nef Genes

David J Nolan; Susanna L Lamers; Rebecca Rose; James J Dollar; Marco Salemi; Michael S McGrath

doi:10.21769/BioProtoc.2334

Single Genome Sequencing of Expressed and Proviral HIV-1 Envelope Glycoprotein 120 (gp120) and nef Genes

Bio Protoc. 2017 Jun 20;7(12):e2334. doi: 10.21769/BioProtoc.2334.

Authors

David J Nolan^{1

2}, Susanna L Lamers¹, Rebecca Rose¹, James J Dollar^{1

2}, Marco Salemi², Michael S McGrath^{3

4}

Affiliations

¹ Bioinfoexperts, LLC, Thibodaux, Louisiana, USA.
² Department of Pathology, Immunology and Laboratory Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA.
³ Departments of Laboratory Medicine, Pathology, and Medicine, University of California at San Francisco, San Francisco, California, USA.
⁴ The AIDS and Cancer Specimen Resource, University of California at San Francisco, San Francisco, California, USA.

Abstract

The current study provides detailed protocols utilized to amplify the complete HIV-1 gp120 and nef genes from single copies of expressed or integrated HIV present in fresh-frozen autopsy tissues of patients who died while on combined antiretroviral therapy (cART) with no detectable plasma viral load (pVL) at death ( Lamers et al., 2016a and 2016b; Rose et al., 2016 ). This method optimizes protocols from previous publications ( Palmer et al., 2005 ; Norström et al., 2012 ; Lamers et al., 2015 ; 2016a and 2016b; Rife et al., 2016 ) to produce single distinct PCR products that can be directly sequenced and includes several cost-saving and time-efficient modifications.

Keywords: Gene amplification; HIV-1; Nested PCR; SGS; Single genome sequencing.

Abstract

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