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为建立和完善海洋线虫相近物种的DNA条形码鉴定方法，本研究以深圳福田红树林湿地自由生活的海洋线虫优势属微口线虫属(Terschellingia)为研究对象，在形态分类鉴定基础上，将DNA条形码技术引入海洋线虫形态相似物种的鉴定，研究线粒体细胞色素氧化酶第一亚基(COⅠ)基因、18S核糖体RNA基因(18S rDNA)和28S rDNA三种基因序列片段的物种分类效果。研究共鉴定出微口线虫属4个不同的形态学种，获得其中3个种的DNA序列。18S rDNA及28S rDNA两种条形码所构建的发育树支持将本属划分成6个类群；Kimura 2 parameter(K2P)种内和种间阈值分别为18S rDNA的0%～2.5%和0.4%～13.7%,28S rDNA的0%和20.5%～84.6%。18S rDNA的MN18F-Nem＿18S＿R引物对所扩增的序列最适合作为微口线虫属种类鉴定的DNA条形码，并可用于区分物种复合体。28S rDNA序列虽然能成功扩增，但扩增效率相对较低；COⅠ基因片段无法在所有物种中成功扩增，推测现有引物可能不适合用于本属序列的提取。研究结果表明，DNA条形码可以用于自由生活海洋线虫形态相似物种的识别，但不同的单基因片段对相同物种的鉴定结果有明显差异。
Abstract：In order to establish and improve the DNA barcoding identification method of marine nematode species, the dominant genus of free-living marine nematode, Terschellingia, was selected as the studying object. After morphological classification, DNA barcode was introduced to identify morphologically similar species in this genus. The species identification performance of mitochondrial cytochrome oxidase first subunit(COⅠ) gene, 18S ribosomal RNA gene(rDNA) and 28S rDNA sequences was determined. Four morphologically different species of genus Terschellingia were identified and the DNA sequences of three species were obtained. The phylogenetic tree constructed with 18S rDNA and 28S rDNA supported the division of this genus into 6 taxa. The intraspecific and interspecific threshold of Kimura 2 parameter(K2P) varied between 0% to 2.5% and between 0.4% to 13.7% for 18S rDNA, respectively, while the intraspecific and interspecific threshold was zero and ranged from 20.5% to 84.6% for 28S rDNA, respectively. The MN18F-Nem＿18S＿R primer of 18S rDNA was the most suitable for being used as the DNA barcode of Terschellingia species, which may serve also as the distinguisher of the members of species complexes. The 28S rDNA sequence can be amplified successfully; however, the amplification met a relatively low success rate. COⅠ gene fragment cannot be successfully amplified among all species. We speculated that the primers available currently are not suitable for retrieving DNA sequence from the species in this genus. Our results indicated that DNA barcoding method can be used to identify morphologically similar species of free-living marine nematode, but their performance are different.
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基本信息:
DOI：10.16441/j.cnki.hdxb.20230168
中图分类号:Q959.17
引用信息:
[1]周仁桂,郭玉清,朱慧兰,等.不同DNA条形码在微口线虫属形态相近物种分类上的应用[J].中国海洋大学学报(自然科学版),2024,54(08):52-62.DOI:10.16441/j.cnki.hdxb.20230168.
基金信息:
福建省自然科学基金项目(2022J01324);; 国家自然科学基金项目(31772416)资助~~
2024-07-16
2024-07-16