Direct chloroplast sequencing: comparison of sequencing platforms and analysis tools for whole chloroplast barcoding

PLoS One. 2014 Oct 17;9(10):e110387. doi: 10.1371/journal.pone.0110387. eCollection 2014.

Abstract

Direct sequencing of total plant DNA using next generation sequencing technologies generates a whole chloroplast genome sequence that has the potential to provide a barcode for use in plant and food identification. Advances in DNA sequencing platforms may make this an attractive approach for routine plant identification. The HiSeq (Illumina) and Ion Torrent (Life Technology) sequencing platforms were used to sequence total DNA from rice to identify polymorphisms in the whole chloroplast genome sequence of a wild rice plant relative to cultivated rice (cv. Nipponbare). Consensus chloroplast sequences were produced by mapping sequence reads to the reference rice chloroplast genome or by de novo assembly and mapping of the resulting contigs to the reference sequence. A total of 122 polymorphisms (SNPs and indels) between the wild and cultivated rice chloroplasts were predicted by these different sequencing and analysis methods. Of these, a total of 102 polymorphisms including 90 SNPs were predicted by both platforms. Indels were more variable with different sequencing methods, with almost all discrepancies found in homopolymers. The Ion Torrent platform gave no apparent false SNP but was less reliable for indels. The methods should be suitable for routine barcoding using appropriate combinations of sequencing platform and data analysis.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Contig Mapping
  • DNA Barcoding, Taxonomic / methods*
  • Genome, Chloroplast / genetics*
  • High-Throughput Nucleotide Sequencing / methods*
  • INDEL Mutation / genetics
  • Molecular Sequence Data
  • Oryza / genetics*
  • Polymorphism, Single Nucleotide / genetics

Grants and funding

The research was funded by Rural Industries Research and Development Corporation (Australia). The funders played no role in study design, data collection, decision to publish or preparation of the manuscript.