基于4种线粒体基因序列的广东省南雄市农区小型兽类DNA条形码分析

姜洪雪, 姚丹丹, 林思亮, 冯志勇

doi:10.11853/j.issn.1003.8280.2022.01.009

中国媒介生物学及控制杂志 >

2022 , Vol. 33 >Issue 1: 48 - 53

DOI: https://doi.org/10.11853/j.issn.1003.8280.2022.01.009

实验研究

基于4种线粒体基因序列的广东省南雄市农区小型兽类DNA条形码分析

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广东省农业科学院植物保护研究所, 植物保护新技术重点实验室, 广东广州 510640

姜洪雪,女,博士,助理研究员,从事鼠类生理生化与防控技术研究,E-mail:jianghongxue805@163.com

收稿日期: 2021-05-24

网络出版日期: 2022-02-17

基金资助

植物重大灾害预警共性关键技术研发创新团队项目（2020KJ113）；广东省农业科学院院长基金项目（201932）

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DNA barcoding analysis of small mammals in the agricultural area in Nanxiong of Guangdong province based on four mitochondrial gene sequences

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Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou, Guangdong 510640, China

Received date: 2021-05-24

Online published: 2022-02-17

Supported by

Research and Innovation Team Project of Common Key Technologies for Plant Major Disaster Early Warning (No. 2020KJ113); President Funding of Guangdong Academy of Agricultural Sciences (No. 201932)

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摘要

目的利用4种线粒体基因对广东省南雄市农田害鼠进行鉴定，探讨不同条形码基因应用于小型兽类鉴定的可靠性。方法 2019年在南雄市珠玑镇农田捕获110只鼠形动物样本，对部分样本通过提取基因组DNA、通用引物扩增线粒体细胞色素C氧化酶亚基Ⅰ（COⅠ）、线粒体细胞色素B（Cytb）、16S rRNA和线粒体DNA控制区（D-loop）基因片段及测序，经同源性比对、遗传距离分析和系统发育树构建鉴定鼠种，并与传统形态学鉴定结果进行比较。结果经过4种基因序列条形码分析，样本隶属于2目2科4属6种。其中臭鼩鼱D-loop不能扩增出条带，臭鼩鼱Cytb、黄毛鼠16S rRNA序列不能正确比对。不同鼠种及不同基因种内遗传距离均小于种间遗传距离。系统发育树显示，同一鼠种均聚为一支，支持率在99%以上，6个样本遗传学鉴定结果与形态学鉴定结果不同。结论 DNA条形码技术是传统形态学鉴定的有力补充，实际应用时，可依据数据库的完整性和准确性合理选择不同的条形码基因，并利用多基因相互验证，以保证鉴定结果准确。

关键词： DNA条形码; 小型兽类; 种类鉴定; 线粒体基因

本文引用格式

姜洪雪, 姚丹丹, 林思亮, 冯志勇 . 基于4种线粒体基因序列的广东省南雄市农区小型兽类DNA条形码分析[J]. 中国媒介生物学及控制杂志, 2022 , 33(1) : 48 -53 . DOI: 10.11853/j.issn.1003.8280.2022.01.009

Abstract

Objective To investigate the reliability of different DNA barcoding genes in identifying small mammals by using four mitochondrial genes to identify the species of farmland rodent pests in Nanxiong city of Guangdong province, China. Methods A total of 110 samples of mouse-like rodents were captured in the farmland of Zhuji town in Nanxiong city in 2019. Genomic DNA extraction was performed for some of these samples, and universal primers were used for the amplification and sequencing of cytochrome c oxidase subunit Ⅰ (COⅠ), cytochrome B (Cytb), 16S rRNA, and mitochondrial DNA control region (D-loop) gene fragments. Rodent species was identified based on homology alignment, genetic distance analysis, and phylogenetic tree construction, and the identification results were compared with the results of conventional morphological identification. Results The DNA barcoding analysis of the four gene sequences showed that the samples belonged to 6 species in 4 genera, 2 families, and 2 orders. The D-loop gene bands of Suncus murinus were not amplified, and the Cytb gene sequence of S. murinus and the 16S rRNA gene sequence of Rattus losea were not correctly identified by BLAST. Intraspecific genetic distance was smaller than interspecific genetic distance for different rodent species and genes. The phylogenetic tree showed that individuals of the same rodent species were clustered into one branch, with a support rate of >99%. Genetic identification results were different from morphological identification results for the 6 samples. Conclusion The DNA barcoding technique is a powerful supplement to conventional morphological identification. In practical application, suitable DNA barcode genes should be selected according to the integrity and accuracy of database, and multi-gene mutual validation should be used to ensure the accuracy of identification results.

Key words： DNA barcoding; Small mammal; Species identification; Mitochondrial gene

参考文献

[1] Liu XH. Rodent biology and management:current status,opinion and challenges in China[J]. J Integr Agr,2019,18(4):830-839. DOI:10.1016/S2095-3119(18)61943-4.
[2] Marquez A,Ulivieri T,Benoit E,et al. House mice as a real sanitary threat of human and animal leptospirosis:Proposal for integrated management[J]. BioMed Res Int,2019,2019:3794876. DOI:10.1155/2019/3794876.
[3] Meerburg BG,Singleton GR,Kijlstra A. Rodent-borne diseases and their risks for public health[J]. Crit Rev Microbiol,2009,35(3):221-270. DOI:10.1080/10408410902989837.
[4] 刘维俊,徐国英,肖方震,等. DNA条形码技术在福建省鼠种鉴定中的应用[J]. 中国媒介生物学及控制杂志,2020,31(2):175-179. DOI:10.11853/j.issn.1003.8280.2020.02.011. Liu WJ,Xu GY,Xiao FZ,et al. Application of DNA barcoding in identifying rodent species in Fujian province,China[J]. Chin J Vector Biol Control,2020,31(2):175-179. DOI:10.11853/j.issn.1003.8280.2020.02.011.(in Chinese)
[5] 肖金花,肖晖,黄大卫. 生物分类学的新动向:DNA条形编码[J]. 动物学报,2004,50(5):852-855. Xiao JH,Xiao H,Huang DW. DNA barcoding:New approach of biological taxonomy[J]. Acta Zool Sinica,2004,50(5):852-855. (in Chinese)
[6] Hebert PDN,Cywinska A,Ball SL,et al. Biological identifications through DNA barcodes[J]. Proc Roy Soc B:Biol Sci,2003,270(1512):313-321. DOI:10.1098/rspb.2002.2218.
[7] Chen WT,Ma XH,Shen YJ,et al. The fish diversity in the upper reaches of the Salween River,Nujiang River,revealed by DNA barcoding[J]. Sci Rep,2015,5:17437. DOI:10.1038/srep17437.
[8] Lu L,Chesters D,Zhang W,et al. Small mammal investigation in spotted fever focus with DNA-barcoding and taxonomic implications on rodents species from Hainan of China[J]. PLoS One,2012,7(8):e43479. DOI:10.1371/journal.pone.0043479.
[9] Wang Y,Wen HY,Zhai DD,et al. DNA barcoding for identification of fishes in Xiangjiaba reservoir area in the downstream section of the Jinsha River[J]. Conserv Genet Resour,2021,13(2):201-208. DOI:10.1007/s12686-021-01196-6.
[10] Nicolas V,Schaeffer B,Missoup AD,et al. Assessment of three mitochondrial genes (16S,Cytb,CO1) for identifying species in the praomyini tribe (Rodentia:Muridae)[J]. PLoS One,2012,7(5):e36586. DOI:10.1371/journal.pone.0036586.
[11] 胡群,马思杰,裘炯良. 4种DNA条形码在黄毛鼠种类鉴定中的比较[J]. 中国媒介生物学及控制杂志,2015,26(3):286-289. DOI:10.11853/j.issn.1003.4692.2015.03.017. Hu Q,Ma SJ,Qiu JL. Comparison of four DNA barcoding in Rattus losea identification[J]. Chin J Vector Biol Control,2015,26(3):286-289. DOI:10.11853/j.issn.1003.4692.2015.03.017.(in Chinese)
[12] 刘蓉蓉,葛德燕,鲁亮,等. 中国姬鼠属种类的DNA条形码鉴定及其分布[J]. 中国媒介生物学及控制杂志,2017,28(2):97-103. DOI:10.11853/j.issn.1003.8280.2017.02.001. Liu RR,Ge DY,Lu L,et al. Identification and distribution of Apodemus species with DNA barcoding in China[J]. Chin J Vector Biol Control,2017,28(2):97-103. DOI:10.11853/j.issn.1003.8280.2017.02.001.(in Chinese)
[13] Liu SY,He K,Chen SD,et al. How many species of Apodemus and Rattus occur in China? A survey based on mitochondrial Cytb and morphological analyses[J]. Zool Res,2018,39(5):309-320. DOI:10.24272/j.issn.2095-8137.2018.053.
[14] 王缠,潘永泰,王静,等. 高原鼢鼠DNA条形码筛选[J]. 草业科学,2020,37(12):2574-2583. DOI:10.11829/j.issn.1001-0629.2020-0054. Wang C,Pan YT,Wang J,et al. Selection of plateau zokor (Eospalax baileyi) DNA barcodes[J]. Pratacult Sci,2020,37(12):2574-2583. DOI:10.11829/j.issn.1001-0629.2020-0054.(in Chinese)
[15] 郑智民,姜志宽,陈安国. 啮齿动物学[M]. 2版. 上海:上海交通大学出版社,2008:62-82. Zheng ZM,Jiang ZK,Chen AG. Rodentology[M]. 2nd ed. Shanghai:Shanghai Jiao Tong University Press,2008:62-82. (in Chinese)
[16] Robins JH,Hingston M,Matisoo-Smith E,et al. Identifying Rattus species using mitochondrial DNA[J]. Mol Ecol Notes,2007,7(5):717-729. DOI:10.1111/j.1471-8286.2007.01752.x.
[17] Ivanova NV,Zemlak TS,Hanner RH,et al. Universal primer cocktails for fish DNA barcoding[J]. Mol Ecol Notes,2007,7(4):544-548. DOI:10.1111/j.1471-8286.2007.01748.x.
[18] 郭丽民, 席进孝, 盖永志, 等. 甘肃省鼠疫疫源地啮齿动物DNA条形码分析[J]. 中国媒介生物学及控制杂志, 2019, 30(2):176-179. DOI:10.11853/j.issn.1003.8280.2019.02.014. Guo LM, Xi JX, Gai YZ, et al. Application of DNA barcoding in the identification of rodent species in plague foci in Gansu province, China[J]. Chin J Vector Biol Control, 2019, 30(2):176-179. DOI:10.11853/j.issn.1003.8280.2019.02.014.(in Chinese)
[19] Tamura K,Peterson D,Peterson N,et al. MEGA 5:Molecular evolutionary genetics analysis using maximum likelihood,evolutionary distance,and maximum parsimony methods[J]. Mol Biol Evol,2011,28(10):2731-2739. DOI:10.1093/molbev/msr121.
[20] Kimura M. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences[J]. J Mol Evol,1980,16(2):111-120. DOI:10.1007/BF01731581.
[21] 何锴,王文智,李权,等. DNA条形码技术在小型兽类鉴定中的探索:以甘肃莲花山为例[J]. 生物多样性,2013,21(2):197-205. DOI:10.3724/SP.J.1003.2013.09160. He K,Wang WZ,Li Q,et al. DNA barcoding in surveys of small mammal community:A case study in Lianhuashan,Gansu province,China[J]. Biodivers Sci,2013,21(2):197-205. DOI:10.3724/SP.J.1003.2013.09160.(in Chinese)
[22] 李争光,丛林,王大伟,等. 黑龙江地区农田害鼠发生特点及DNA条形码鉴定技术的应用[J]. 植物保护,2018,44(6):145-151,167. DOI:10.16688/j.zwbh.2018001. Li ZG,Cong L,Wang DW,et al. Species identification of main pest rodents by DNA barcoding and their occurrence characteristics from croplands in Heilongjiang province[J]. Plant Prot,2018,44(6):145-151,167. DOI:10.16688/j.zwbh.2018001. (in Chinese)
[23] 陈宝宝,孙养信,范锁平,等. DNA条形码技术在陕西省鼠形动物鉴定中的应用[J]. 中国人兽共患病学报,2020,36(4):325-329,339. DOI:10.3969/j.issn.1002-2694.2020.00.038. Chen BB,Sun YX,Fan SP,et al. Application for identification of murine-like animals by DNA barcoding in Shaanxi province[J]. Chin J Zoonoses,2020,36(4):325-329,339. DOI:10.3969/j.issn.1002-2694.2020.00.038.(in Chinese)
[24] Li J,Zheng X,Cai YS,et al. DNA barcoding of Murinae (Rodentia:Muridae) and Arvicolinae (Rodentia:Cricetidae) distributed in China[J]. Mol Ecol Resour,2015,15(1):153-167. DOI:10.1111/1755-0998.12279.
[25] 彭培英,郭宪国. 卡氏小鼠的研究动态[J]. 医学动物防制,2014,30(3):290-292. DOI:10.7629/yxdwfz201403018. Peng PY,Guo XG. Research status on the field mouse,Mus caroli[J]. J Med Pest Control,2014,30(3):290-292. DOI:10.7629/yxdwfz201403018.(in Chinese)
[26] Hebert PDN,Ratnasingham S,de Waard JR. Barcoding animal life:cytochrome c oxidase subunit 1 divergences among closely related species[J]. Proc Roy Soc B:Biol Sci,2003,270 Suppl 1:S96-99. DOI:10.1098/rsbl.2003.0025.
[27] Tobe SS,Kitchener AC,Linacre AMT. Reconstructing mammalian phylogenies:A detailed comparison of the cytochrome b and cytochrome oxidase subunit I mitochondrial genes[J]. PLoS One,2010,5(11):e14156. DOI:10.1371/journal.pone. 0014156.
[28] 刘山林. DNA条形码参考数据集构建和序列分析相关的新兴技术[J]. 生物多样性,2019,27(5):526-533. DOI:10.17520/biods.2018209. Liu SL. DNA barcoding and emerging reference construction and data analysis technologies[J]. Biodivers Sci,2019,27(5):526-533. DOI:10.17520/biods.2018209.(in Chinese)

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