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Abstract
Although DNA barcoding has become a useful tool for species identification and biodiversity surveys in plant sciences, there remains little consensus concerning appropriate sampling strategies and the treatment of indels. To address these two issues, we sampled 39 populations for nine Taxus species across their entire ranges, with two to three individuals per population randomly sampled. We sequenced one core DNA barcode (matK) and three supplementary regions (trnH-psbA, trnL-trnF and ITS) for all samples to test the effects of sampling design and the utility of indels. Our results suggested that increasing sampling within-population did not change the clustering of individuals, and that meant within-population P-distances were zero for most populations in all regions. Based on the markers tested here, comparison of methods either including or excluding indels indicated that discrimination and nodal support of monophyletic groups were significantly increased when indels were included. Thus we concluded that one individual per population was adequate to represent the within-population variation in these species for DNA barcoding, and that intra-specific sampling was best focused on representing the entire ranges of certain taxa. We also found that indels occurring in the chloroplast trnL-trnF and trnH-psbA regions were informative to differentiate among for closely related taxa barcoding, and we proposed that indel-coding methods should be considered for use in future for closed related plant species DNA barcoding projects on or below generic level.
Highlights
DNA barcoding is a technique to identify species by using standardized DNA sequences [1]
Length variation was observed in the trnL-trnF, trnH-psbA and internal transcribed spacer (ITS) sequences (Table 1), which ranged from 797 bp to 852 bp, 532 bp to 981 bp and 1135 bp to 1141 bp, respectively
The number of individuals required to create a reliable reference for valid species identification has been one of the basic issues considered since the beginning of the DNA barcoding initiative [18,21]
Summary
DNA barcoding is a technique to identify species by using standardized DNA sequences [1]. The mitochondrial cytochrome oxidase subunit I gene (COI) is a widely used barcoding region in a range of animal groups, but is unsuitable for plant barcoding due to its low substitution rate and frequent intra-molecular recombination of the mitochondrial genome in land plants [6,7,8]. Many studies have already compared the performance of a range of candidate DNA loci as barcodes in different plant taxa (e.g., [6,7,9]), the optimal choice of DNA regions adopted for plant barcoding has not yet achieved consensus [8,10,11]. Based on a comprehensive evaluation of seven candidate DNA regions on a large dataset, the Consortium for the Barcode of Life (CBOL) Plant
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