Evaluating sampling strategy for DNA barcoding study of coastal and inland halo-tolerant Poaceae and Chenopodiaceae: A case study for increased sample size

Environmental conditions in coastal salt marsh habitats have led to the development of specialist genetic adaptations. We evaluated six DNA barcode loci of the 53 species of Poaceae and 15 species of Chenopodiaceae from China's coastal salt marsh area and inland area. Our results indicate that the optimum DNA barcode was ITS for coastal salt-tolerant Poaceae and matK for the Chenopodiaceae. Sampling strategies for ten common species of Poaceae and Chenopodiaceae were analyzed according to optimum barcode. We found that by increasing the number of samples collected from the coastal salt marsh area on the basis of inland samples, the number of haplotypes of Arundinella hirta, Digitaria ciliaris, Eleusine indica, Imperata cylindrica, Setaria viridis, and Chenopodium glaucum increased, with a principal coordinate plot clearly showing increased distribution points. The results of a Mann-Whitney test showed that for Digitaria ciliaris, Eleusine indica, Imperata cylindrica, and Setaria viridis, the distribution of intraspecific genetic distances was significantly different when samples from the coastal salt marsh area were included (P < 0.01). These results suggest that increasing the sample size in specialist habitats can improve measurements of intraspecific genetic diversity, and will have a positive effect on the application of the DNA barcodes in widely distributed species. The results of random sampling showed that when sample size reached 11 for Chloris virgata, Chenopodium glaucum, and Dysphania ambrosioides, 13 for Setaria viridis, and 15 for Eleusine indica, Imperata cylindrica and Chenopodium album, average intraspecific distance tended to reach stability. These results indicate that the sample size for DNA barcode of globally distributed species should be increased to 11–15.

滨海盐沼生境的环境条件，促使物种演化出特化的遗传适应机制。本研究针对采自中国滨海盐沼区与内陆区域的53种禾本科（Poaceae）植物及15种藜科（Chenopodiaceae）植物，对6个DNA条形码（DNA barcode）位点开展评估。研究结果显示，耐滨海盐渍的禾本科植物的最优DNA条形码为ITS，而藜科植物的最优DNA条形码则为matK。基于最优条形码，本研究对10种常见禾本科与藜科植物的采样策略进行了分析。研究发现，在以内陆样本为基础的前提下，增加滨海盐沼区的采样量后，野古草（Arundinella hirta）、马唐（Digitaria ciliaris）、牛筋草（Eleusine indica）、白茅（Imperata cylindrica）、狗尾草（Setaria viridis）以及灰绿藜（Chenopodium glaucum）的单倍型数量均有所提升；主坐标图（principal coordinate plot）亦清晰显示其分布点数量增加。曼-惠特尼检验（Mann-Whitney test）结果表明，对于马唐、牛筋草、白茅与狗尾草而言，当纳入滨海盐沼区样本后，其种内遗传距离的分布存在显著差异（P < 0.01）。上述结果表明，在特化生境中增加采样量，可提升种内遗传多样性的检测精度，同时对DNA条形码在广布物种中的应用具有积极作用。随机采样结果显示，对于虎尾草（Chloris virgata）、灰绿藜及腺毛藜（Dysphania ambrosioides），当采样量达到11份时；对于狗尾草，采样量达到13份时；对于牛筋草、白茅以及藜（Chenopodium album），采样量达到15份时，其平均种内遗传距离均趋于稳定。上述结果提示，广布物种DNA条形码研究的采样量应提升至11~15份。