Successful Recovery of Nuclear Protein-Coding Genes from Small Insects in Museums Using Illumina Sequencing Public Deposited

http://ir.library.oregonstate.edu/concern/articles/wm117q84z

This is the publisher’s final pdf. The published article is copyrighted by the author(s) and published by the Public Library of Science. The published article can be found at:  http://www.plosone.org/

Data Availability Statement: Raw reads for all museum and reference specimens are submitted to NCBI Sequence Read Archive (accessions SRR2939013– SRR2939027). Focal gene fragments recovered from the de novo assembly of Lagriinae n. gen. and those that were newly sequenced for the phylogeny of Lagriinae are deposited in GenBank (accessions KU233685-KU234083). Focal gene fragments from PCR/Sanger sequencing and the IlluminaMerged sequences of carabids are also deposited in GenBank (accessions KU233685- KU234083). The Tribolium castaneum and Bembidion sp. nr transversale query sequences used to probe our museum specimens for the 67 nuclear protein-coding gene fragments and all alignments used in phylogenetic analyses (including the DeNovo, FarRef, and NearRef sequences), as well as trees from the phylogenetic tests, are deposited in Dryad (data available from the Dryad Digital Repository:  http://doi.org/xx).

Supporting information is available online at:  http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0143929#sec051

Descriptions

Attribute NameValues
Creator
Abstract or Summary
  • In this paper we explore high-throughput Illumina sequencing of nuclear protein-coding, ribosomal, and mitochondrial genes in small, dried insects stored in natural history collections. We sequenced one tenebrionid beetle and 12 carabid beetles ranging in size from 3.7 to 9.7 mm in length that have been stored in various museums for 4 to 84 years. Although we chose a number of old, small specimens for which we expected low sequence recovery, we successfully recovered at least some low-copy nuclear protein-coding genes from all specimens. For example, in one 56-year-old beetle, 4.4 mm in length, our de novo assembly recovered about 63% of approximately 41,900 nucleotides in a target suite of 67 nuclear protein-coding gene fragments, and 70% using a reference-based assembly. Even in the least successfully sequenced carabid specimen, reference-based assembly yielded fragments that were at least 50% of the target length for 34 of 67 nuclear protein-coding gene fragments. Exploration of alternative references for reference-based assembly revealed few signs of bias created by the reference. For all specimens we recovered almost complete copies of ribosomal and mitochondrial genes. We verified the general accuracy of the sequences through comparisons with sequences obtained from PCR and Sanger sequencing, including of conspecific, fresh specimens, and through phylogenetic analysis that tested the placement of sequences in predicted regions. A few possible inaccuracies in the sequences were detected, but these rarely affected the phylogenetic placement of the samples. Although our sample sizes are low, an exploratory regression study suggests that the dominant factor in predicting success at recovering nuclear protein-coding genes is a high number of Illumina reads, with success at PCR of COI and killing by immersion in ethanol being secondary factors; in analyses of only high-read samples, the primary significant explanatory variable was body length, with small beetles being more successfully sequenced.
Resource Type
DOI
Date Available
Date Issued
Citation
  • Kanda, K., Pflug, J. M., Sproul, J. S., Dasenko, M. A., & Maddison, D. R. (2015). Successful Recovery of Nuclear Protein-Coding Genes from Small Insects in Museums Using Illumina Sequencing. PLoS ONE, 10(12), e0143929. doi:10.1371/journal.pone.0143929
Series
Rights Statement
Funding Statement (additional comments about funding)
Publisher
Peer Reviewed
Language
Replaces
Additional Information
  • description.provenance : Approved for entry into archive by Patricia Black(patricia.black@oregonstate.edu) on 2016-02-04T20:26:04Z (GMT) No. of bitstreams: 2 license_rdf: 1370 bytes, checksum: cd1af5ab51bcc7a5280cf305303530e9 (MD5) KandaKojunIntegrativeBioSuccessfulRecoverNuclear.pdf: 6988858 bytes, checksum: 8a730720bee6252816def5881a7fd8e3 (MD5)
  • description.provenance : Made available in DSpace on 2016-02-04T20:26:04Z (GMT). No. of bitstreams: 2 license_rdf: 1370 bytes, checksum: cd1af5ab51bcc7a5280cf305303530e9 (MD5) KandaKojunIntegrativeBioSuccessfulRecoverNuclear.pdf: 6988858 bytes, checksum: 8a730720bee6252816def5881a7fd8e3 (MD5) Previous issue date: 2015-12-30
  • description.provenance : Submitted by Patricia Black (patricia.black@oregonstate.edu) on 2016-02-04T20:25:25Z No. of bitstreams: 2 license_rdf: 1370 bytes, checksum: cd1af5ab51bcc7a5280cf305303530e9 (MD5) KandaKojunIntegrativeBioSuccessfulRecoverNuclear.pdf: 6988858 bytes, checksum: 8a730720bee6252816def5881a7fd8e3 (MD5)

Relationships

Parents:

This work has no parents.

Last modified

Downloadable Content

Download PDF

Items