Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers
Next generation sequencing (NGS) platforms have the ability to generate almost limitless numbers of sequence reads starting with a PCR product. This gives the illusion that it is possible to analyze minor variants in a viral population. However, including a PCR step obscures the sampling depth of th...
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| Format: | Article |
| Language: | English |
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MDPI AG
2020-08-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/12/8/850 |
| _version_ | 1797560411081408512 |
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| author | Shuntai Zhou Ronald Swanstrom |
| author_facet | Shuntai Zhou Ronald Swanstrom |
| author_sort | Shuntai Zhou |
| collection | DOAJ |
| description | Next generation sequencing (NGS) platforms have the ability to generate almost limitless numbers of sequence reads starting with a PCR product. This gives the illusion that it is possible to analyze minor variants in a viral population. However, including a PCR step obscures the sampling depth of the viral population, the key parameter needed to understand the utility of the data set for finding minor variants. Also, these high throughput sequencing platforms are error prone at the level where minor variants are of interest, confounding the interpretation of detected minor variants. A simple strategy has been applied in multiple applications of NGS to solve these problems. Prior to PCR, individual molecules are “tagged” with a unique molecular identifier (UMI) that can be used to establish the actual sample size of viral genomes sequenced after PCR and sequencing. In addition, since PCR generates many copies of each sequence tagged to a specific UMI, a template consensus sequence (TCS) can be created from the many reads of each template, removing virtually all of the method error. From this perspective we examine our own use of a UMI, called Primer ID, in the detection of minor drug resistant variants in HIV-1 populations. |
| first_indexed | 2024-03-10T17:59:22Z |
| format | Article |
| id | doaj.art-8a0500fb622f4640aab87b89998f1de6 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-10T17:59:22Z |
| publishDate | 2020-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-8a0500fb622f4640aab87b89998f1de62023-11-20T09:00:09ZengMDPI AGViruses1999-49152020-08-0112885010.3390/v12080850Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular IdentifiersShuntai Zhou0Ronald Swanstrom1Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USALineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USANext generation sequencing (NGS) platforms have the ability to generate almost limitless numbers of sequence reads starting with a PCR product. This gives the illusion that it is possible to analyze minor variants in a viral population. However, including a PCR step obscures the sampling depth of the viral population, the key parameter needed to understand the utility of the data set for finding minor variants. Also, these high throughput sequencing platforms are error prone at the level where minor variants are of interest, confounding the interpretation of detected minor variants. A simple strategy has been applied in multiple applications of NGS to solve these problems. Prior to PCR, individual molecules are “tagged” with a unique molecular identifier (UMI) that can be used to establish the actual sample size of viral genomes sequenced after PCR and sequencing. In addition, since PCR generates many copies of each sequence tagged to a specific UMI, a template consensus sequence (TCS) can be created from the many reads of each template, removing virtually all of the method error. From this perspective we examine our own use of a UMI, called Primer ID, in the detection of minor drug resistant variants in HIV-1 populations.https://www.mdpi.com/1999-4915/12/8/850drug resistance mutationsunique molecular identifierprimer IDnext generation sequencingHIV |
| spellingShingle | Shuntai Zhou Ronald Swanstrom Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers Viruses drug resistance mutations unique molecular identifier primer ID next generation sequencing HIV |
| title | Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers |
| title_full | Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers |
| title_fullStr | Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers |
| title_full_unstemmed | Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers |
| title_short | Fact and Fiction about 1%: Next Generation Sequencing and the Detection of Minor Drug Resistant Variants in HIV-1 Populations with and without Unique Molecular Identifiers |
| title_sort | fact and fiction about 1 next generation sequencing and the detection of minor drug resistant variants in hiv 1 populations with and without unique molecular identifiers |
| topic | drug resistance mutations unique molecular identifier primer ID next generation sequencing HIV |
| url | https://www.mdpi.com/1999-4915/12/8/850 |
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