BMC Genomics (Dec 2020)

Efficient COI barcoding using high throughput single-end 400 bp sequencing

  • Chentao Yang,
  • Yuxuan Zheng,
  • Shangjin Tan,
  • Guanliang Meng,
  • Wei Rao,
  • Caiqing Yang,
  • David G. Bourne,
  • Paul A. O’Brien,
  • Junqiang Xu,
  • Sha Liao,
  • Ao Chen,
  • Xiaowei Chen,
  • Xinrui Jia,
  • Ai-bing Zhang,
  • Shanlin Liu

DOI
https://doi.org/10.1186/s12864-020-07255-w
Journal volume & issue
Vol. 21, no. 1
pp. 1 – 10

Abstract

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Abstract Background Over the last decade, the rapid development of high-throughput sequencing platforms has accelerated species description and assisted morphological classification through DNA barcoding. However, the current high-throughput DNA barcoding methods cannot obtain full-length barcode sequences due to read length limitations (e.g. a maximum read length of 300 bp for the Illumina’s MiSeq system), or are hindered by a relatively high cost or low sequencing output (e.g. a maximum number of eight million reads per cell for the PacBio’s SEQUEL II system). Results Pooled cytochrome c oxidase subunit I (COI) barcodes from individual specimens were sequenced on the MGISEQ-2000 platform using the single-end 400 bp (SE400) module. We present a bioinformatic pipeline, HIFI-SE, that takes reads generated from the 5′ and 3′ ends of the COI barcode region and assembles them into full-length barcodes. HIFI-SE is written in Python and includes four function modules of filter, assign, assembly and taxonomy. We applied the HIFI-SE to a set of 845 samples (30 marine invertebrates, 815 insects) and delivered a total of 747 fully assembled COI barcodes as well as 70 Wolbachia and fungi symbionts. Compared to their corresponding Sanger sequences (72 sequences available), nearly all samples (71/72) were correctly and accurately assembled, including 46 samples that had a similarity score of 100% and 25 of ca. 99%. Conclusions The HIFI-SE pipeline represents an efficient way to produce standard full-length barcodes, while the reasonable cost and high sensitivity of our method can contribute considerably more DNA barcodes under the same budget. Our method thereby advances DNA-based species identification from diverse ecosystems and increases the number of relevant applications.

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