DNA条形码技术在大型红藻分子鉴定中的应用

Application of DNA Barcoding Technology in Molecular Identification of Red Macroalgae

本研究对浙江省枸杞岛贻贝养殖区大型海藻资源进行了调查，在形态学观察的基础上，选取93株红藻样品运用DNA分子条形码技术进行分子鉴定分析。采用的3个分子标记分别为rbcL（叶绿体基因组中1，5-二磷酸核酮糖羧化／加氧酶大亚基基因），COI（细胞色C氧化酶亚基I）和UPA（23 SrRNA基因片段，通用质体扩增区1基因片段。通过实验，我们共扩增出rbcL基因片段79条，扩增成功率为84．95％；共扩增出COI基因片段63条，扩增成功率为67．74％；共扩增出UPA基因片段76条，扩增成功率为81．72％。3个分子标记的测序成功率分别为：rbcL基因78．49％，COI基因29．03％，UPA基因72．04％。COI基因的扩增效率和测序成功率均低于rbcL和UPA基因。基于3个基因构建（Neighbor-joining，N—J）系统进化树表明，rbcL基因可以将红藻样品归类为5目，6科，7属，12种；COI基因可以将样品归类为4目，5科，5属，5种；UPA基因可以将样品归类为6目，7科，8属，11种。rbcL、COI和UPA3个基因结合可以将样品归类为7目8科11属21种。研究表明rbcL基因以及UPA基因较适合做为鉴定大型红藻的分子标记，C01基因做为红藻分子标记的潜力有待进一步开发和研究。

This study investigated macroalgae resources in the mussel culture zone of Gouqi island, Zhejiang province. 93 samples of red algae based on the morphological observation were selected and analyzed by DNA molecular barcoding technique for molecular identification. Three used molecular markers were rbcL （1,5-ribulose diphosphate carboxylase/oxygenase large-subunit gene in the chloroplast genome）, CO1 （cytochrome C oxidase subunit I） and UPA （23S rRNA gene fragment, universal plastid amplification region） gene fragments. Through experiments, 79 rbcL gene fragments were amplified with a success rate of 84.95%; 63 COI gene fragments were amplified with a success rate of 67.74%; 76 UPA gene fragments were amplified with a success rate of 81.72%. The sequencing success rate of rbcL gene, COI gene and UPA gene were 78.49%, 29.03% and 72.04% respectively. The amplification efficiency and sequencing success rate of COI gene were lower than those ofrbcL and UPA genes. Neighbor-joining （NJ） phylogenetic tree based on three genes showed that rbcL gene could classify the samples into 5 orders, 6 families, 7 genera and 12 species. COIgene could classify the samples into 4 orders, 5 families, 5 genera and 5 species. UPA gene could classify the samples into 6 orders, 7 families, 8 genera and 11 species. However, the combination ofrbcL, COI and UPA genes could classify the samples into 7 orders, 8 families, 11 genera and 21 species. The results showed that rbcL and UPA genes were more suitable as molecular markers for the identification of red macroalgae but the potential of COI gene as a molecular marker of red macroalgae remained to be further developed and studied.