Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome
Abstract Background With a whole genome duplication event and wealth of biological data, salmonids are excellent model organisms for studying evolutionary processes, fates of duplicated genes and genetic and physiological processes associated with complex behavioral phenotypes. It is surprising ther...
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ftbiomed:oai:biomedcentral.com:1471-2164-9-404 2023-05-15T15:31:29+02:00 Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome Quinn, Nicole L Levenkova, Natasha Chow, William Bouffard, Pascal Boroevich, Keith A Knight, James R Jarvie, Thomas P Lubieniecki, Krzysztof P Desany, Brian A Koop, Ben F Harkins, Timothy T Davidson, William S 2008-08-28 http://www.biomedcentral.com/1471-2164/9/404 en eng BioMed Central Ltd. http://www.biomedcentral.com/1471-2164/9/404 Copyright 2008 Quinn et al; licensee BioMed Central Ltd. Research article 2008 ftbiomed 2008-09-26T23:12:44Z Abstract Background With a whole genome duplication event and wealth of biological data, salmonids are excellent model organisms for studying evolutionary processes, fates of duplicated genes and genetic and physiological processes associated with complex behavioral phenotypes. It is surprising therefore, that no salmonid genome has been sequenced. Atlantic salmon ( Salmo salar ) is a good representative salmonid for sequencing given its importance in aquaculture and the genomic resources available. However, the size and complexity of the genome combined with the lack of a sequenced reference genome from a closely related fish makes assembly challenging. Given the cost and time limitations of Sanger sequencing as well as recent improvements to next generation sequencing technologies, we examined the feasibility of using the Genome Sequencer (GS) FLX pyrosequencing system to obtain the sequence of a salmonid genome. Eight pooled BACs belonging to a minimum tiling path covering ~1 Mb of the Atlantic salmon genome were sequenced by GS FLX shotgun and Long Paired End sequencing and compared with a ninth BAC sequenced by Sanger sequencing of a shotgun library. Results An initial assembly using only GS FLX shotgun sequences (average read length 248.5 bp) with ~30× coverage allowed gene identification, but was incomplete even when 126 Sanger-generated BAC-end sequences (~0.09× coverage) were incorporated. The addition of paired end sequencing reads (additional ~26× coverage) produced a final assembly comprising 175 contigs assembled into four scaffolds with 171 gaps. Sanger sequencing of the ninth BAC (~10.5× coverage) produced nine contigs and two scaffolds. The number of scaffolds produced by the GS FLX assembly was comparable to Sanger-generated sequencing; however, the number of gaps was much higher in the GS FLX assembly. Conclusion These results represent the first use of GS FLX paired end reads for de novo sequence assembly. Our data demonstrated that this improved the GS FLX assemblies; however, with respect to de novo sequencing of complex genomes, the GS FLX technology is limited to gene mining and establishing a set of ordered sequence contigs. Currently, for a salmonid reference sequence, it appears that a substantial portion of sequencing should be done using Sanger technology. Article in Journal/Newspaper Atlantic salmon Salmo salar BioMed Central |
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English |
description |
Abstract Background With a whole genome duplication event and wealth of biological data, salmonids are excellent model organisms for studying evolutionary processes, fates of duplicated genes and genetic and physiological processes associated with complex behavioral phenotypes. It is surprising therefore, that no salmonid genome has been sequenced. Atlantic salmon ( Salmo salar ) is a good representative salmonid for sequencing given its importance in aquaculture and the genomic resources available. However, the size and complexity of the genome combined with the lack of a sequenced reference genome from a closely related fish makes assembly challenging. Given the cost and time limitations of Sanger sequencing as well as recent improvements to next generation sequencing technologies, we examined the feasibility of using the Genome Sequencer (GS) FLX pyrosequencing system to obtain the sequence of a salmonid genome. Eight pooled BACs belonging to a minimum tiling path covering ~1 Mb of the Atlantic salmon genome were sequenced by GS FLX shotgun and Long Paired End sequencing and compared with a ninth BAC sequenced by Sanger sequencing of a shotgun library. Results An initial assembly using only GS FLX shotgun sequences (average read length 248.5 bp) with ~30× coverage allowed gene identification, but was incomplete even when 126 Sanger-generated BAC-end sequences (~0.09× coverage) were incorporated. The addition of paired end sequencing reads (additional ~26× coverage) produced a final assembly comprising 175 contigs assembled into four scaffolds with 171 gaps. Sanger sequencing of the ninth BAC (~10.5× coverage) produced nine contigs and two scaffolds. The number of scaffolds produced by the GS FLX assembly was comparable to Sanger-generated sequencing; however, the number of gaps was much higher in the GS FLX assembly. Conclusion These results represent the first use of GS FLX paired end reads for de novo sequence assembly. Our data demonstrated that this improved the GS FLX assemblies; however, with respect to de novo sequencing of complex genomes, the GS FLX technology is limited to gene mining and establishing a set of ordered sequence contigs. Currently, for a salmonid reference sequence, it appears that a substantial portion of sequencing should be done using Sanger technology. |
format |
Article in Journal/Newspaper |
author |
Quinn, Nicole L Levenkova, Natasha Chow, William Bouffard, Pascal Boroevich, Keith A Knight, James R Jarvie, Thomas P Lubieniecki, Krzysztof P Desany, Brian A Koop, Ben F Harkins, Timothy T Davidson, William S |
spellingShingle |
Quinn, Nicole L Levenkova, Natasha Chow, William Bouffard, Pascal Boroevich, Keith A Knight, James R Jarvie, Thomas P Lubieniecki, Krzysztof P Desany, Brian A Koop, Ben F Harkins, Timothy T Davidson, William S Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
author_facet |
Quinn, Nicole L Levenkova, Natasha Chow, William Bouffard, Pascal Boroevich, Keith A Knight, James R Jarvie, Thomas P Lubieniecki, Krzysztof P Desany, Brian A Koop, Ben F Harkins, Timothy T Davidson, William S |
author_sort |
Quinn, Nicole L |
title |
Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
title_short |
Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
title_full |
Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
title_fullStr |
Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
title_full_unstemmed |
Assessing the feasibility of GS FLX Pyrosequencing for sequencing the Atlantic salmon genome |
title_sort |
assessing the feasibility of gs flx pyrosequencing for sequencing the atlantic salmon genome |
publisher |
BioMed Central Ltd. |
publishDate |
2008 |
url |
http://www.biomedcentral.com/1471-2164/9/404 |
genre |
Atlantic salmon Salmo salar |
genre_facet |
Atlantic salmon Salmo salar |
op_relation |
http://www.biomedcentral.com/1471-2164/9/404 |
op_rights |
Copyright 2008 Quinn et al; licensee BioMed Central Ltd. |
_version_ |
1766362000265314304 |