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Comparison of a high-resolution melting assay to next-generation sequencing for analysis of HIV diversity.
- Source :
-
Journal of clinical microbiology [J Clin Microbiol] 2012 Sep; Vol. 50 (9), pp. 3054-9. Date of Electronic Publication: 2012 Jul 11. - Publication Year :
- 2012
-
Abstract
- Next-generation sequencing (NGS) has recently been used for analysis of HIV diversity, but this method is labor-intensive, costly, and requires complex protocols for data analysis. We compared diversity measures obtained using NGS data to those obtained using a diversity assay based on high-resolution melting (HRM) of DNA duplexes. The HRM diversity assay provides a single numeric score that reflects the level of diversity in the region analyzed. HIV gag and env from individuals in Rakai, Uganda, were analyzed in a previous study using NGS (n = 220 samples from 110 individuals). Three sequence-based diversity measures were calculated from the NGS sequence data (percent diversity, percent complexity, and Shannon entropy). The amplicon pools used for NGS were analyzed with the HRM diversity assay. HRM scores were significantly associated with sequence-based measures of HIV diversity for both gag and env (P < 0.001 for all measures). The level of diversity measured by the HRM diversity assay and NGS increased over time in both regions analyzed (P < 0.001 for all measures except for percent complexity in gag), and similar amounts of diversification were observed with both methods (P < 0.001 for all measures except for percent complexity in gag). Diversity measures obtained using the HRM diversity assay were significantly associated with those from NGS, and similar increases in diversity over time were detected by both methods. The HRM diversity assay is faster and less expensive than NGS, facilitating rapid analysis of large studies of HIV diversity and evolution.
- Subjects :
- Adult
HIV isolation & purification
Humans
Molecular Biology methods
Nucleic Acid Denaturation
Uganda
Virology methods
env Gene Products, Human Immunodeficiency Virus genetics
gag Gene Products, Human Immunodeficiency Virus genetics
DNA, Viral genetics
Genetic Variation
HIV classification
HIV genetics
HIV Infections virology
High-Throughput Nucleotide Sequencing methods
Transition Temperature
Subjects
Details
- Language :
- English
- ISSN :
- 1098-660X
- Volume :
- 50
- Issue :
- 9
- Database :
- MEDLINE
- Journal :
- Journal of clinical microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 22785188
- Full Text :
- https://doi.org/10.1128/JCM.01460-12