HIV-1 RNA was extracted using the QIAamp Viral RNA Mini Kit (Qiagen, US) from 140 μl of patients’ plasma. The cDNA was generated using the SuperScript® III RT enzyme (Invitrogen, Life Technologies, MA, USA) with gene specific primer 6231R, as described by us (Thermo Scientific) [15 (link)]. The first-round PCR was performed using the high-fidelity KAPA HiFiHotStart Ready Mix (2x) (KAPA Biosystem, MA, USA) with primers 0682F and 6231R primers. The second-round PCR was performed using 0702F-BssHII and 5798R-SalI which has BssHII and SalI restriction sites, respectively. The amplified product was restriction digested followed by gel purification using the QIAquickGel Extraction Kit (Qiagen, USA). The gag-pol fragment (HXB2:0702-5798) was cloned in pNL43Δgag-pol plasmid following digestion with BssHII and SalI (New England Biolab, US) and ligation with T4 DNA ligase (New England Biolab, US). The recombinant viruses were produced by transfecting the plasmids into the 293T cell line using FuGENE® HD Transfection Reagent (Promega, US). All the molecular clones were sequenced bi-directionally. No primary DRM was observed in any of the clones. M50I was present in seven of the sequences from the HIV-1B (n=3) and the HIV-1C (n=4) strains.