CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing
Background Long-read nanopore sequencing technology is of particular significance for taxonomic identification at or below the species level. For many environmental samples, the total extractable DNA is far below the current input requirements of nanopore sequencing, preventing “sam...
| Main Authors: | , , , , |
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| Other Authors: | |
| Format: | Article |
| Language: | English |
| Published: |
BioMed Central
2018
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| Online Access: | http://hdl.handle.net/1721.1/114584 https://orcid.org/0000-0003-4547-4747 https://orcid.org/0000-0003-2652-8017 |
| _version_ | 1811071358934712320 |
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| author | Hachey, Julie Ruvkun, Gary Mojarro, Angel Zuber, Maria Carr, Christopher E. |
| author2 | Massachusetts Institute of Technology. Department of Earth, Atmospheric, and Planetary Sciences |
| author_facet | Massachusetts Institute of Technology. Department of Earth, Atmospheric, and Planetary Sciences Hachey, Julie Ruvkun, Gary Mojarro, Angel Zuber, Maria Carr, Christopher E. |
| author_sort | Hachey, Julie |
| collection | MIT |
| description | Background
Long-read nanopore sequencing technology is of particular significance for taxonomic identification at or below the species level. For many environmental samples, the total extractable DNA is far below the current input requirements of nanopore sequencing, preventing “sample to sequence” metagenomics from low-biomass or recalcitrant samples. Results
Here we address this problem by employing carrier sequencing, a method to sequence low-input DNA by preparing the target DNA with a genomic carrier to achieve ideal library preparation and sequencing stoichiometry without amplification. We then use CarrierSeq, a sequence analysis workflow to identify the low-input target reads from the genomic carrier. We tested CarrierSeq experimentally by sequencing from a combination of 0.2 ng Bacillus subtilis ATCC 6633 DNA in a background of 1000 ng Enterobacteria phage λ DNA. After filtering of carrier, low quality, and low complexity reads, we detected target reads (B. subtilis), contamination reads, and “high quality noise reads” (HQNRs) not mapping to the carrier, target or known lab contaminants. These reads appear to be artifacts of the nanopore sequencing process as they are associated with specific channels (pores).
Conclusion
By treating sequencing as a Poisson arrival process, we implement a statistical test to reject data from channels dominated by HQNRs while retaining low-input target reads. |
| first_indexed | 2024-09-23T08:49:58Z |
| format | Article |
| id | mit-1721.1/114584 |
| institution | Massachusetts Institute of Technology |
| language | English |
| last_indexed | 2024-09-23T08:49:58Z |
| publishDate | 2018 |
| publisher | BioMed Central |
| record_format | dspace |
| spelling | mit-1721.1/1145842024-05-15T02:15:50Z CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing Hachey, Julie Ruvkun, Gary Mojarro, Angel Zuber, Maria Carr, Christopher E. Massachusetts Institute of Technology. Department of Earth, Atmospheric, and Planetary Sciences Mojarro, Angel Zuber, Maria Carr, Christopher E Background Long-read nanopore sequencing technology is of particular significance for taxonomic identification at or below the species level. For many environmental samples, the total extractable DNA is far below the current input requirements of nanopore sequencing, preventing “sample to sequence” metagenomics from low-biomass or recalcitrant samples. Results Here we address this problem by employing carrier sequencing, a method to sequence low-input DNA by preparing the target DNA with a genomic carrier to achieve ideal library preparation and sequencing stoichiometry without amplification. We then use CarrierSeq, a sequence analysis workflow to identify the low-input target reads from the genomic carrier. We tested CarrierSeq experimentally by sequencing from a combination of 0.2 ng Bacillus subtilis ATCC 6633 DNA in a background of 1000 ng Enterobacteria phage λ DNA. After filtering of carrier, low quality, and low complexity reads, we detected target reads (B. subtilis), contamination reads, and “high quality noise reads” (HQNRs) not mapping to the carrier, target or known lab contaminants. These reads appear to be artifacts of the nanopore sequencing process as they are associated with specific channels (pores). Conclusion By treating sequencing as a Poisson arrival process, we implement a statistical test to reject data from channels dominated by HQNRs while retaining low-input target reads. United States. National Aeronautics and Space Administration (Award NNX15AF85G) 2018-04-06T14:14:46Z 2018-04-06T14:14:46Z 2018-03 2017-10 2018-04-01T12:59:11Z Article http://purl.org/eprint/type/JournalArticle 1471-2105 http://hdl.handle.net/1721.1/114584 Mojarro, Angel et al. "CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing." BMC Bioinformatics 19 (March 2018):108 © 2018 The Authors https://orcid.org/0000-0003-4547-4747 https://orcid.org/0000-0003-2652-8017 en http://dx.doi.org/10.1186/s12859-018-2124-3 BMC Bioinformatics Creative Commons Attribution http://creativecommons.org/licenses/by/4.0/ The Author(s). application/pdf BioMed Central BioMed Central |
| spellingShingle | Hachey, Julie Ruvkun, Gary Mojarro, Angel Zuber, Maria Carr, Christopher E. CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title | CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title_full | CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title_fullStr | CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title_full_unstemmed | CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title_short | CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing |
| title_sort | carrierseq a sequence analysis workflow for low input nanopore sequencing |
| url | http://hdl.handle.net/1721.1/114584 https://orcid.org/0000-0003-4547-4747 https://orcid.org/0000-0003-2652-8017 |
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