The Internal Transcribed Spacer (ITS) Region and <em>trnhH-psbA</em> Are Suitable Candidate Loci for DNA Barcoding of Tropical Tree Species of India

Background DNA barcoding as a tool for species identification has been successful in animals and other organisms, including certain groups of plants. The exploration of this new tool for species identification, particularly in tree species, is very scanty from biodiversity-rich countries like India. rbcL and matK are standard barcode loci while ITS, and trnH-psbA are considered as supplementary loci for plants. Methodology and Principal Findings Plant barcode loci, namely, rbcL, matK, ITS, trnH-psbA, and the recently proposed ITS2, were tested for their efficacy as barcode loci using 300 accessions of tropical tree species. We tested these loci for PCR, sequencing success, and species discrimination ability using three methods. rbcL was the best locus as far as PCR and sequencing success rate were concerned, but not for the species discrimination ability of tropical tree species. ITS and trnH-psbA were the second best loci in PCR and sequencing success, respectively. The species discrimination ability of ITS ranged from 24.4 percent to 74.3 percent and that of trnH-psbA was 25.6 percent to 67.7 percent, depending upon the data set and the method used. matK provided the least PCR success, followed by ITS2 (59. 0%). Species resolution by ITS2 and rbcL ranged from 9.0 percent to 48.7 percent and 13.2 percent to 43.6 percent, respectively. Further, we observed that the NCBI nucleotide database is poorly represented by the sequences of barcode loci studied here for tree species. Conclusion Although a conservative approach of a success rate of 60–70 percent by both ITS and trnH-psbA may not be considered as highly successful but would certainly help in large-scale biodiversity inventorization, particularly for tropical tree species, considering the standard success rate of plant DNA barcode program reported so far. The recommended matK and rbcL primers combination may not work in tropical tree species as barcode markers.背景 DNA条形码（DNA barcoding）作为物种鉴定工具，已在动物及包括部分植物类群在内的其他生物中取得成功。但在生物多样性丰富的国家（如印度），针对该工具在物种鉴定（尤其是林木树种）中的应用探索仍十分匮乏。rbcL与matK为植物标准条形码基因座，而ITS、trnH-psbA则被视为植物辅助条形码基因座。 研究方法与核心发现 本研究以300份热带树种种质材料为研究对象，对rbcL、matK、ITS、trnH-psbA以及新近提出的ITS2等植物条形码基因座的鉴定效能进行了检测。本研究通过三种方法评估了上述基因座的PCR扩增成功率、测序成功率与物种区分能力。就PCR与测序成功率而言，rbcL为表现最优的基因座，但在热带树种的物种区分能力上却并非最佳。ITS与trnH-psbA则分别在PCR扩增成功率与测序成功率中位列第二。ITS的物种区分能力介于24.4%至74.3%之间，trnH-psbA的物种区分能力则为25.6%至67.7%，具体数值取决于所采用的数据集与分析方法。matK的PCR扩增成功率最低，其次为ITS2（59.0%）。ITS2与rbcL的物种分辨率分别介于9.0%至48.7%、13.2%至43.6%之间。此外，本研究发现，NCBI核苷酸数据库中针对林木树种的上述研究所用条形码基因座序列资源十分匮乏。 结论 综合目前已报道的植物DNA条形码项目标准成功率来看，尽管ITS与trnH-psbA的成功率处于60%~70%的保守区间，尚未达到极高成功水平，但仍可对大规模生物多样性清查（尤其是热带树种的清查）提供有力助力。目前推荐的matK与rbcL引物组合，或无法作为条形码标记应用于热带树种的物种鉴定。