对锦葵科植物样品的ITS、ITS2、rbcL、matK和psbA-trnH序列进行PCR扩增和测序, 比较各序列的扩增效率、测序成功率、种内和种间变异的差异以及barcoding gap图, 使用BLAST1和Nearest Distance方法评价不同序列的鉴定能力, 进而从这些候选序列中筛选出较适合锦葵科植物鉴别的DNA条形码序列。结果表明, ITS序列在采集的锦葵科植物11个种26个样品中的扩增成功率较高, 其种内、种间变异差异和barcoding gap较ITS2、psbA-trnH及rbcL序列具有更明显的优势, 且纳入60个属316个种共1 228个样品的网上数据后, 其鉴定成功率可达89.9%。psbA-trnH序列的扩增和测序成功率最高, 其鉴定成功率为63.2%, 并能鉴别一些ITS序列无法鉴别的种。实验结果表明, ITS和psbA-trnH是较适合鉴别锦葵科植物的DNA条形码序列组合。
To screen the best candidate sequence that can be used as a universal DNA barcode to identify species in Malvaceae, we amplified by PCR and sequenced the DNA fragments of 5 hot regions (ITS, ITS2, rbcL, matK and psbA-trnH) in samples from the family Malvaceae. PCR amplification and sequencing, differential intra- and interspecific divergences, DNA barcoding gap and identification (using BLAST1 and nearest distance methods) were used to evaluate
the discrimination ability of these candidate sequences. The ITS region had fairly high amplification efficiency in 26 plant samples belonging to 11 Malvaceae species and better intra- and interspecific divergences and DNA barcoding gap than any other region tested (ITS2, psbA-trnH and rbcL). In addition, the rate of successful identification with the ITS region was 89.9% at the species level for after added 1 228 plant samples from the GenBank database of Malvaceae belonging to 316 species from 60 distinct genera. The psbA-trnH region had the best amplification efficiency and sequencing efficiency; the rate of successful identification with the psbA-trnH region was 63.2%, and the region can discriminate some species that cannot be successfully discriminated by the ITS region. We propose that ITS and psbA-trnH are a promising DNA barcode sequence combination for identification.
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