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目的:研究DNA条形码技术在稻田小型害虫稻蓟马、稻叶蝉和稻飞虱种类鉴别中的适用性。方法:利用DNA条形码的通用引物对稻田3种稻蓟马、5种稻叶蝉和3种稻飞虱的线粒体细胞色素C氧化酶亚基I(Cytochrome c oxidase subunit I, COI)基因序列进行扩增、测序和分析，以Kimura-2-parameter (K2P)模型计算种间和种内遗传距离，并以邻接法(Neighbor-Joining, NJ)构建系统发育树。结果:通过PCR扩增测序结合GenBank公共序列筛选获得11种水稻害虫COI序列共34条，其序列长度位于646～658 bp之间，且碱基组成存在明显的A+T碱基偏向性。K2P遗传距离分析结果显示，11种水稻害虫的种内和种间平均遗传距离分别为0.002和0.326,最大种内和最小种间遗传距离分别为0.004和0.091,种内与种间遗传距离之间存在显著的条形码间隙(barcode gap)。系统发育分析结果显示，NJ进化树中同一物种昆虫聚集于同一进化分支，聚类分析与形态学鉴定结果一致。结论:基于COI基因的DNA条形码技术可以准确鉴别我国水稻上三类小型害虫——稻蓟马、稻叶蝉和稻飞虱。
Abstract：Aims: This paper aims to test the feasibility of DNA barcoding technology for species identification of thrips(Thysanoptera), leafhoppers and planthoppers(Hemiptera) from rice paddies. Methods: The mitochondrial cytochrome oxidase subunit I(COI) fragments were amplified using DNA barcoding universal primers from 3 species of rice thrips, 5 species of rice leafhoppers and 3 species of rice planthoppers, and then sequenced for DNA barcoding analysis. The intra-and inter-species genetic distances were estimated based on the Kimura-2-parameter(K2P) model. A Neighbor-Joining(NJ) tree was constructed using the K2P model in the MEGA version 11.0 software. Results: A total of 34 COI sequences were obtained from 11 species of rice pests by PCR amplification and GenBank screening, which ranged from 646 bp to 658 bp in length and showed an obvious preference to A+T bases. Genetic distance analysis revealed that the mean intraspecific genetic distance among the 11 pest species was 0.326, while the mean interspecific genetic distance was 0.002. The maximum intraspecific and minimum interspecific genetic distances were 0.004 and 0.091, respectively. A barcode gap was obviously observed between the intraspecific and interspecific genetic distances. The phylogenetic result revealed that each of the species was clustered in a respective branch in the NJ tree, consistent with those results based on morphological characteristics. Conclusions: COI can be effective as a suitable DNA barcode to identify three kinds of small-sized pests(thrips, leafhoppers and planthoppers) from rice paddies in China..
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基本信息:
DOI：
中图分类号:S435.112
引用信息:
[1]程依情,邵珠龙,王正亮等.基于DNA条形码的稻蓟马、稻叶蝉和稻飞虱种类鉴别[J].中国计量大学学报,2023,34(01):44-50.
基金信息:
国家自然科学基金项目(NO.31972347,U21A20223); 浙江省属高校基本科研业务费专项项目(NO.2022YW82)