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J Virol Methods. 2010 Sep;168(1-2):114-20. doi: 10.1016/j.jviromet.2010.04.030. Epub 2010 May 6.

Comparison of standard PCR/cloning to single genome sequencing for analysis of HIV-1 populations.

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Tufts University School of Medicine, Tufts Medical Center, Boston, MA 02111, USA.


To compare standard PCR/cloning and single genome sequencing (SGS) in their ability to reflect actual intra-patient polymorphism of HIV-1 populations, a total of 530 HIV-1 pro-pol sequences obtained by both sequencing techniques from a set of 17 ART naïve patient specimens was analyzed. For each specimen, 12 and 15 sequences, on average, were characterized by the two techniques. Using phylogenetic analysis, tests for panmixia and entropy, and Bland-Altman plots, no difference in population structure or genetic diversity was shown in 14 of the 17 subjects. Evidence of sampling bias by the presence of subsets of identical sequences was found by either method. Overall, the study shows that neither method was more biased than the other, and providing that an adequate number of PCR templates is analyzed, and that the bulk sequencing captures the diversity of the viral population, either method is likely to provide a similar measure of population diversity.

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