From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research
Next-generation sequencing (NGS) applications have flourished in the last decade, permitting the identification of cancer driver genes and profoundly expanding the possibilities of genomic studies of cancer, including melanoma. Here we aimed to present a technical review across many of the methodolo...
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-11-01
|
| Series: | Life |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2075-1729/12/11/1939 |
| _version_ | 1797464819010371584 |
|---|---|
| author | Adrián Muñoz-Barrera Luis A. Rubio-Rodríguez Ana Díaz-de Usera David Jáspez José M. Lorenzo-Salazar Rafaela González-Montelongo Víctor García-Olivares Carlos Flores |
| author_facet | Adrián Muñoz-Barrera Luis A. Rubio-Rodríguez Ana Díaz-de Usera David Jáspez José M. Lorenzo-Salazar Rafaela González-Montelongo Víctor García-Olivares Carlos Flores |
| author_sort | Adrián Muñoz-Barrera |
| collection | DOAJ |
| description | Next-generation sequencing (NGS) applications have flourished in the last decade, permitting the identification of cancer driver genes and profoundly expanding the possibilities of genomic studies of cancer, including melanoma. Here we aimed to present a technical review across many of the methodological approaches brought by the use of NGS applications with a focus on assessing germline and somatic sequence variation. We provide cautionary notes and discuss key technical details involved in library preparation, the most common problems with the samples, and guidance to circumvent them. We also provide an overview of the sequence-based methods for cancer genomics, exposing the pros and cons of targeted sequencing vs. exome or whole-genome sequencing (WGS), the fundamentals of the most common commercial platforms, and a comparison of throughputs and key applications. Details of the steps and the main software involved in the bioinformatics processing of the sequencing results, from preprocessing to variant prioritization and filtering, are also provided in the context of the full spectrum of genetic variation (SNVs, indels, CNVs, structural variation, and gene fusions). Finally, we put the emphasis on selected bioinformatic pipelines behind (a) short-read WGS identification of small germline and somatic variants, (b) detection of gene fusions from transcriptomes, and (c) de novo assembly of genomes from long-read WGS data. Overall, we provide comprehensive guidance across the main methodological procedures involved in obtaining sequencing results for the most common short- and long-read NGS platforms, highlighting key applications in melanoma research. |
| first_indexed | 2024-03-09T18:12:33Z |
| format | Article |
| id | doaj.art-2b0f87de69ac440cb0852a59149a79ad |
| institution | Directory Open Access Journal |
| issn | 2075-1729 |
| language | English |
| last_indexed | 2024-03-09T18:12:33Z |
| publishDate | 2022-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Life |
| spelling | doaj.art-2b0f87de69ac440cb0852a59149a79ad2023-11-24T08:58:23ZengMDPI AGLife2075-17292022-11-011211193910.3390/life12111939From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma ResearchAdrián Muñoz-Barrera0Luis A. Rubio-Rodríguez1Ana Díaz-de Usera2David Jáspez3José M. Lorenzo-Salazar4Rafaela González-Montelongo5Víctor García-Olivares6Carlos Flores7Genomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainGenomics Division, Instituto Tecnológico y de Energías Renovables (ITER), 38600 Santa Cruz de Tenerife, SpainNext-generation sequencing (NGS) applications have flourished in the last decade, permitting the identification of cancer driver genes and profoundly expanding the possibilities of genomic studies of cancer, including melanoma. Here we aimed to present a technical review across many of the methodological approaches brought by the use of NGS applications with a focus on assessing germline and somatic sequence variation. We provide cautionary notes and discuss key technical details involved in library preparation, the most common problems with the samples, and guidance to circumvent them. We also provide an overview of the sequence-based methods for cancer genomics, exposing the pros and cons of targeted sequencing vs. exome or whole-genome sequencing (WGS), the fundamentals of the most common commercial platforms, and a comparison of throughputs and key applications. Details of the steps and the main software involved in the bioinformatics processing of the sequencing results, from preprocessing to variant prioritization and filtering, are also provided in the context of the full spectrum of genetic variation (SNVs, indels, CNVs, structural variation, and gene fusions). Finally, we put the emphasis on selected bioinformatic pipelines behind (a) short-read WGS identification of small germline and somatic variants, (b) detection of gene fusions from transcriptomes, and (c) de novo assembly of genomes from long-read WGS data. Overall, we provide comprehensive guidance across the main methodological procedures involved in obtaining sequencing results for the most common short- and long-read NGS platforms, highlighting key applications in melanoma research.https://www.mdpi.com/2075-1729/12/11/1939cancer genomicsmelanomanext-generation sequencingthird-generation sequencingnanoporebioinformatic workflows |
| spellingShingle | Adrián Muñoz-Barrera Luis A. Rubio-Rodríguez Ana Díaz-de Usera David Jáspez José M. Lorenzo-Salazar Rafaela González-Montelongo Víctor García-Olivares Carlos Flores From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research Life cancer genomics melanoma next-generation sequencing third-generation sequencing nanopore bioinformatic workflows |
| title | From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research |
| title_full | From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research |
| title_fullStr | From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research |
| title_full_unstemmed | From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research |
| title_short | From Samples to Germline and Somatic Sequence Variation: A Focus on Next-Generation Sequencing in Melanoma Research |
| title_sort | from samples to germline and somatic sequence variation a focus on next generation sequencing in melanoma research |
| topic | cancer genomics melanoma next-generation sequencing third-generation sequencing nanopore bioinformatic workflows |
| url | https://www.mdpi.com/2075-1729/12/11/1939 |
| work_keys_str_mv | AT adrianmunozbarrera fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT luisarubiorodriguez fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT anadiazdeusera fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT davidjaspez fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT josemlorenzosalazar fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT rafaelagonzalezmontelongo fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT victorgarciaolivares fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch AT carlosflores fromsamplestogermlineandsomaticsequencevariationafocusonnextgenerationsequencinginmelanomaresearch |