{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,21]],"date-time":"2026-08-21T12:55:42Z","timestamp":1787316942215,"version":"build-2736575974"},"reference-count":32,"publisher":"SAGE Publications","issue":"11","license":[{"start":{"date-parts":[[2024,11,1]],"date-time":"2024-11-01T00:00:00Z","timestamp":1730419200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.sagepub.com\/page\/policies\/text-and-data-mining-license"}],"content-domain":{"domain":["journals.sagepub.com"],"crossmark-restriction":true},"short-container-title":["AIDS Research and Human Retroviruses"],"published-print":{"date-parts":[[2024,11]]},"abstract":"<jats:p>\n                    Our goal was to assess the accuracy of next generation sequencing (NGS) compared with Sanger. We performed single genome amplification (SGA) of HIV-1\n                    <jats:italic toggle=\"yes\">gp160<\/jats:italic>\n                    on extracted tissue DNA from two HIV+ individuals. Amplicons (\n                    <jats:italic toggle=\"yes\">n<\/jats:italic>\n                    = 30) were sequenced with Sanger or reamplified with barcoded primers and pooled before sequencing using Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PB). For each amplicon, a consensus sequence for NGS reads was obtained by (1) mapping reads to the Sanger sequence when available (\u201creference-based\u201d) or (2) mapping reads to a \u201cpseudo-reference\u201d sequence, i.e., a consensus sequence of a subset of NGS reads (\u201creference-free\u201d). PB reads were clustered based on genetic similarity. A Sanger consensus sequence was obtained for 23\/30 amplicons, for which all NGS consensus sequences were identical (\n                    <jats:italic toggle=\"yes\">n<\/jats:italic>\n                    = 9) or nearly identical (\n                    <jats:italic toggle=\"yes\">n<\/jats:italic>\n                    = 14) compared with Sanger. For the nine mismatches between Sanger\/NGS, the nucleotide in the NGS sequence matched all other sequences from that patient. Of the 7\/30 amplicons without a Sanger sequence, NGS sequences had \u226535 ambiguous calls in five amplicons and 0 ambiguities in two amplicons. Analysis of the electropherograms showed failure of a single sequencing primer for the latter two amplicons (consistent with a single template) and overlapping peaks for the other five (consistent with multiple templates). Clustering results closely followed the Sanger\/NGS consensus results, where amplicons derived from a single template also had a single cluster and vice versa (with one exception, which could be the result of barcode misidentification). Representative sequences from the clusters contained 2\u201313 differences compared with Sanger\/NGS. In summary, we show that both ONT and PB can produce amplicon consensus sequences with similar or higher accuracy compared with Sanger and, importantly, without the need for a known reference sequence. Clustering could be useful in some circumstances to predict or confirm the presence of multiple starting templates.\n                  <\/jats:p>","DOI":"10.1089\/aid.2024.0012","type":"journal-article","created":{"date-parts":[[2024,7,3]],"date-time":"2024-07-03T10:12:33Z","timestamp":1720001553000},"page":"659-669","update-policy":"https:\/\/doi.org\/10.1177\/sage-journals-update-policy","source":"Crossref","is-referenced-by-count":2,"title":["Comparing Gold-Standard Sanger Sequencing with Two Next-Generation Sequencing Platforms of HIV-1\n                    <i>gp160<\/i>\n                    Single Genome Amplicons"],"prefix":"10.1177","volume":"40","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-9114-7002","authenticated-orcid":false,"given":"David J.","family":"Nolan","sequence":"first","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jonathan","family":"DaRoza","sequence":"additional","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robin","family":"Brody","sequence":"additional","affiliation":[{"name":"Molecular Medicine, UMass Chan Medical School, Worcester, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Krishna","family":"Ganta","sequence":"additional","affiliation":[{"name":"Molecular Medicine, UMass Chan Medical School, Worcester, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katherine","family":"Luzuriaga","sequence":"additional","affiliation":[{"name":"Molecular Medicine, UMass Chan Medical School, Worcester, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chris","family":"Huston","sequence":"additional","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Simon","family":"Rosenthal","sequence":"additional","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Susanna L.","family":"Lamers","sequence":"additional","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5059-9176","authenticated-orcid":false,"given":"Rebecca","family":"Rose","sequence":"additional","affiliation":[{"name":"Bioinfoexperts LLC, Thibodaux, Louisiana, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"179","published-online":{"date-parts":[[2024,11,6]]},"reference":[{"key":"e_1_3_5_2_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/18.7.1687"},{"key":"e_1_3_5_3_2","doi-asserted-by":"publisher","DOI":"10.1128\/jvi.64.2.864-872.1990"},{"key":"e_1_3_5_4_2","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.64.12.5840-5850.1990"},{"key":"e_1_3_5_5_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.43.1.406-413.2005"},{"key":"e_1_3_5_6_2","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.02660-07"},{"key":"e_1_3_5_7_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jviromet.2009.08.002"},{"key":"e_1_3_5_8_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.74.12.5463"},{"key":"e_1_3_5_9_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12864-018-4611-3"},{"key":"e_1_3_5_10_2","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2015.249623"},{"key":"e_1_3_5_11_2","doi-asserted-by":"publisher","DOI":"10.1007\/s11908-014-0401-5"},{"key":"e_1_3_5_12_2","doi-asserted-by":"publisher","DOI":"10.1172\/JCI124291"},{"key":"e_1_3_5_13_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2017.09.081"},{"key":"e_1_3_5_14_2","doi-asserted-by":"publisher","DOI":"10.3390\/cells10102577"},{"key":"e_1_3_5_15_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1008853"},{"key":"e_1_3_5_16_2","doi-asserted-by":"publisher","DOI":"10.1128\/jvi.00543-23"},{"key":"e_1_3_5_17_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkv630"},{"key":"e_1_3_5_18_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz657"},{"key":"e_1_3_5_19_2","doi-asserted-by":"publisher","DOI":"10.1093\/ve\/vey007"},{"key":"e_1_3_5_20_2","doi-asserted-by":"publisher","DOI":"10.1186\/s13059-020-1935-5"},{"key":"e_1_3_5_21_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkq543"},{"key":"e_1_3_5_22_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-022-01539-7"},{"key":"e_1_3_5_23_2","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.00674-16"},{"key":"e_1_3_5_24_2","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.00684-16"},{"key":"e_1_3_5_25_2","doi-asserted-by":"publisher","DOI":"10.21769\/BioProtoc.2334"},{"key":"e_1_3_5_26_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1000080107"},{"key":"e_1_3_5_27_2","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/bty149"},{"key":"e_1_3_5_28_2","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btw152"},{"key":"e_1_3_5_29_2","doi-asserted-by":"publisher","DOI":"10.1093\/molbev\/mst010"},{"key":"e_1_3_5_30_2","doi-asserted-by":"publisher","DOI":"10.7717\/peerj.12983"},{"key":"e_1_3_5_31_2","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btp352"},{"key":"e_1_3_5_32_2","doi-asserted-by":"publisher","DOI":"10.7717\/peerj.2584"},{"key":"e_1_3_5_33_2","doi-asserted-by":"publisher","DOI":"10.1093\/molbev\/msaa015"}],"container-title":["AIDS 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