Mutation detection with next-generation resequencing through a mediator genome.
The affordability of next generation sequencing (NGS) is transforming the field of mutation analysis in bacteria. The genetic basis for phenotype alteration can be identified directly by sequencing the entire genome of the mutant and comparing it to the wild-type (WT) genome, thus identifying acquir...
Saved in:
| Main Authors: | , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2010-12-01
|
| Series: | PLoS ONE |
| Online Access: | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0015628&type=printable |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1849695051018403840 |
|---|---|
| author | Omri Wurtzel Mally Dori-Bachash Shmuel Pietrokovski Edouard Jurkevitch Rotem Sorek |
| author_facet | Omri Wurtzel Mally Dori-Bachash Shmuel Pietrokovski Edouard Jurkevitch Rotem Sorek |
| author_sort | Omri Wurtzel |
| collection | DOAJ |
| description | The affordability of next generation sequencing (NGS) is transforming the field of mutation analysis in bacteria. The genetic basis for phenotype alteration can be identified directly by sequencing the entire genome of the mutant and comparing it to the wild-type (WT) genome, thus identifying acquired mutations. A major limitation for this approach is the need for an a-priori sequenced reference genome for the WT organism, as the short reads of most current NGS approaches usually prohibit de-novo genome assembly. To overcome this limitation we propose a general framework that utilizes the genome of relative organisms as mediators for comparing WT and mutant bacteria. Under this framework, both mutant and WT genomes are sequenced with NGS, and the short sequencing reads are mapped to the mediator genome. Variations between the mutant and the mediator that recur in the WT are ignored, thus pinpointing the differences between the mutant and the WT. To validate this approach we sequenced the genome of Bdellovibrio bacteriovorus 109J, an obligatory bacterial predator, and its prey-independent mutant, and compared both to the mediator species Bdellovibrio bacteriovorus HD100. Although the mutant and the mediator sequences differed in more than 28,000 nucleotide positions, our approach enabled pinpointing the single causative mutation. Experimental validation in 53 additional mutants further established the implicated gene. Our approach extends the applicability of NGS-based mutant analyses beyond the domain of available reference genomes. |
| format | Article |
| id | doaj-art-4081f2695dc843b68db6cf8aaa8464e6 |
| institution | DOAJ |
| issn | 1932-6203 |
| language | English |
| publishDate | 2010-12-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj-art-4081f2695dc843b68db6cf8aaa8464e62025-08-20T03:19:53ZengPublic Library of Science (PLoS)PLoS ONE1932-62032010-12-01512e1562810.1371/journal.pone.0015628Mutation detection with next-generation resequencing through a mediator genome.Omri WurtzelMally Dori-BachashShmuel PietrokovskiEdouard JurkevitchRotem SorekThe affordability of next generation sequencing (NGS) is transforming the field of mutation analysis in bacteria. The genetic basis for phenotype alteration can be identified directly by sequencing the entire genome of the mutant and comparing it to the wild-type (WT) genome, thus identifying acquired mutations. A major limitation for this approach is the need for an a-priori sequenced reference genome for the WT organism, as the short reads of most current NGS approaches usually prohibit de-novo genome assembly. To overcome this limitation we propose a general framework that utilizes the genome of relative organisms as mediators for comparing WT and mutant bacteria. Under this framework, both mutant and WT genomes are sequenced with NGS, and the short sequencing reads are mapped to the mediator genome. Variations between the mutant and the mediator that recur in the WT are ignored, thus pinpointing the differences between the mutant and the WT. To validate this approach we sequenced the genome of Bdellovibrio bacteriovorus 109J, an obligatory bacterial predator, and its prey-independent mutant, and compared both to the mediator species Bdellovibrio bacteriovorus HD100. Although the mutant and the mediator sequences differed in more than 28,000 nucleotide positions, our approach enabled pinpointing the single causative mutation. Experimental validation in 53 additional mutants further established the implicated gene. Our approach extends the applicability of NGS-based mutant analyses beyond the domain of available reference genomes.https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0015628&type=printable |
| spellingShingle | Omri Wurtzel Mally Dori-Bachash Shmuel Pietrokovski Edouard Jurkevitch Rotem Sorek Mutation detection with next-generation resequencing through a mediator genome. PLoS ONE |
| title | Mutation detection with next-generation resequencing through a mediator genome. |
| title_full | Mutation detection with next-generation resequencing through a mediator genome. |
| title_fullStr | Mutation detection with next-generation resequencing through a mediator genome. |
| title_full_unstemmed | Mutation detection with next-generation resequencing through a mediator genome. |
| title_short | Mutation detection with next-generation resequencing through a mediator genome. |
| title_sort | mutation detection with next generation resequencing through a mediator genome |
| url | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0015628&type=printable |
| work_keys_str_mv | AT omriwurtzel mutationdetectionwithnextgenerationresequencingthroughamediatorgenome AT mallydoribachash mutationdetectionwithnextgenerationresequencingthroughamediatorgenome AT shmuelpietrokovski mutationdetectionwithnextgenerationresequencingthroughamediatorgenome AT edouardjurkevitch mutationdetectionwithnextgenerationresequencingthroughamediatorgenome AT rotemsorek mutationdetectionwithnextgenerationresequencingthroughamediatorgenome |