基于DNA条形码技术构建王朗国家级自然保护区陆生脊椎动物遗传资源数据库及物种鉴定

doi:10.17520/biods.2022661

生物多样性 ›› 2023, Vol. 31 ›› Issue (7): 22661. DOI: 10.17520/biods.2022661

• 研究报告: 动物多样性 • 上一篇下一篇

基于DNA条形码技术构建王朗国家级自然保护区陆生脊椎动物遗传资源数据库及物种鉴定

邢超¹^,², 林依²^,³, 周智强⁴, 赵联军⁴, 蒋仕伟⁵, 林蓁蓁²^,^*(), 徐基良¹^,^*(), 詹祥江²

1.北京林业大学生态与自然保护学院, 北京 100083
2.中国科学院动物研究所动物生态与保护生物学重点实验室, 北京 100101
3.中国科学院大学, 北京 100049
4.四川王朗国家级自然保护区管理局, 四川平武 622550
5.平武县林业和草原局, 四川平武 622550

收稿日期:2022-11-25 接受日期:2023-04-14 出版日期:2023-07-20 发布日期:2023-04-25
通讯作者: *E-mail: xujiliang@bjfu.edu.cn; linzz@ioz.ac.cn
作者简介:linzz@ioz.ac.cn
*E-mail: xujiliang@bjfu.edu.cn;
基金资助:
四川王朗国家级自然保护区物种库项目

The establishment of terrestrial vertebrate genetic resource bank and species identification based on DNA barcoding in Wanglang National Nature Reserve

Chao Xing¹^,², Yi Lin²^,³, Zhiqiang Zhou⁴, Lianjun Zhao⁴, Shiwei Jiang⁵, Zhenzhen Lin²^,^*(), Jiliang Xu¹^,^*(), Xiangjiang Zhan²

1. College of Ecology and Nature Conservation, Beijing Forestry University, Beijing 100083
2. Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101
3. University of Chinese Academy of Sciences, Beijing 100049
4. Wanglang National Nature Reserve Administration, Pingwu, Sichuan 622550
5. Pingwu Forestry and Grassland Bureau, Pingwu, Sichuan 622550

Received:2022-11-25 Accepted:2023-04-14 Online:2023-07-20 Published:2023-04-25
Contact: *E-mail: xujiliang@bjfu.edu.cn; linzz@ioz.ac.cn

摘要/Abstract

摘要：

DNA条形码(DNA barcode)是基因组中较短的、种内变异相对稳定的基因序列,已经成为生物多样性保护研究中物种鉴定、生物多样性评估的有力手段之一。四川王朗国家级自然保护区地处青藏高原东缘, 属于世界生物多样性热点地区, 具有丰富的生物资源, 在我国珍稀动物保护领域具有重要地位。目前保护区已累积了大量的陆生脊椎动物监测数据, 但缺乏遗传资源本底调查和基础的遗传资源数据库。本研究基于DNA条形码技术, 以四川王朗国家级自然保护区为主要研究区域, 基于样线法和博物馆标本调研, 对所采集的314份样品进行DNA条形码分析, 共鉴定兽类、鸟类、两栖类18目35科74种, 首次获得了王朗齿突蟾(Scutiger wanglangensis)的线粒体基因(COI、12S-16S、16S、Cytb)及核基因(RAG1)的条形码序列信息, 并通过比较不同监测方法说明了DNA条形码技术在动物多样性调查中的应用前景。本研究基于DNA条形码技术最终获得了216份DNA条形码数据, 初步建立了保护区陆生脊椎动物遗传资源数据库, 该数据库将为评估保护区生物多样性提供基础信息, 为动物保护和管理工作提供技术支持。

关键词: DNA条形码, 王朗国家级自然保护区, 陆生脊椎动物, 生物多样性, 条形码数据库

Abstract

Aims: DNA barcoding using the short, highly conserved regions of the genome to make species-level identifications, is widely used in the species identification and diversity assessment of plants and animals in biodiversity conservation research. Wanglang National Nature Reserve is a global biodiversity hot spot located in the eastern part of Qinghai-Tibet Plateau with high level of biodiversity and has a leading role in the wildlife conservation in China. Although a lot of survey data of species diversity have been collected, genetic resource of terrestrial vertebrates in the reserve remains largely unclear. To address this, our study aims to establish a DNA barcoding database for terrestrial vertebrate species for the reserve.

Methods: We collected genetic samples mainly using non-invasive sampling along the line transects in Wanglang National Nature Reserve and from museum specimens. We extracted DNA from the samples to obtain the barcode sequences by PCR. Each DNA barcode sequences were blasted against NCBI database for species identification. For those sequences that we could not directly get the matches in the reference database, we constructed phylogenetic trees using MEGA 11.0 with the aim to identify the species based on the sequences of its relative species from the public database.

Results: The total of 314 samples including tissues from carcasses, feathers, feces and eggshells were collected and 74 species were identified using 16 pairs DNA barcoding primers. The sequences of mitochondrial COI, Cytb, 12S-16S, 16S genes and sequences of nuclear RAG1 gene of Scutiger wanglangensis were firstly reported. Compared with other wildlife monitoring techniques such as camera traps, drift fences and pitfall traps in the same time interval, DNA barcoding is more efficient in all taxon groups and has better performance in bird monitoring. Finally, A terrestrial vertebrate genetic resource bank contains 314 substantial samples and 216 DNA barcodes has been established.

Conclusion: Our study established an elementary DNA barcoding database of 74 terrestrial vertebrate species in the reserve, highlights the utility of DNA barcoding technique in wild animal diversity surveys, and provides a basic catalog for assessing local species diversity to improve conservation research and management in the reserve.

Key words: DNA barcoding, Wanglang National Nature Reserve, terrestrial vertebrates, biodiversity, barcode database

邢超, 林依, 周智强, 赵联军, 蒋仕伟, 林蓁蓁, 徐基良, 詹祥江 (2023) 基于DNA条形码技术构建王朗国家级自然保护区陆生脊椎动物遗传资源数据库及物种鉴定. 生物多样性, 31, 22661. DOI: 10.17520/biods.2022661.

Chao Xing, Yi Lin, Zhiqiang Zhou, Lianjun Zhao, Shiwei Jiang, Zhenzhen Lin, Jiliang Xu, Xiangjiang Zhan (2023) The establishment of terrestrial vertebrate genetic resource bank and species identification based on DNA barcoding in Wanglang National Nature Reserve. Biodiversity Science, 31, 22661. DOI: 10.17520/biods.2022661.

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链接本文: https://www.biodiversity-science.net/CN/10.17520/biods.2022661

https://www.biodiversity-science.net/CN/Y2023/V31/I7/22661

图/表 7

参考文献 79

[1]	Alacs EA, Georges A, FitzSimmons NN, Robertson J (2010) DNA detective: A review of molecular approaches to wildlife forensics. Forensic Science, Medicine, and Pathology, 6, 180-194. DOI PMID
[2]	Cao YZ, Zhao LJ, Zhang TX, Wang C, Zheng Y, Xu L, Ran JH (2022) Daily activity pattern and population density of Tetraophasis obscurus in the Wanglang National Nature Reserve, Sichuan. Sichuan Journal of Zoology, 41, 379-386. (in Chinese with English abstract)
	[曹亚珍, 赵联军, 张塔星, 王灿, 郑勇, 许路, 冉江洪 (2022) 四川王朗国家级自然保护区红喉雉鹑的日活动节律及种群密度. 四川动物, 41, 379-386.]
[3]	Carracedo A, Bär W, Lincoln P, Mayr W, Morling N, Olaisen B, Schneider P, Budowle B, Brinkmann B, Gill P, Holland M, Tully G, Wilson M (2000) DNA commission of the International Society for Forensic Genetics: Guidelines for mitochondrial DNA typing. Forensic Science International, 110, 79-85. PMID
[4]	Chaves PB, Graeff VG, Lion MB, Oliveira LR, Eizirik E (2012) DNA barcoding meets molecular scatology: Short mtDNA sequences for standardized species assignment of carnivore noninvasive samples. Molecular Ecology Resources, 12, 18-35. DOI PMID
[5]	Che J, Jiang K, Yan F, Zhang YP (2020) Amphibians and Reptiles in Tibet—Diversity and Evolution. Science Press, Beijing. (in Chinese)
	[车静, 蒋珂, 颜芳, 张亚平 (2000) 西藏两栖爬行动物——多样性与进化. 科学出版社, 北京.]
[6]	Chen W, Bi K, Fu JZ (2009) Frequent mitochondrial gene introgression among high elevation Tibetan megophryid frogs revealed by conflicting gene genealogies. Molecular Ecology, 18, 2856-2876. DOI PMID
[7]	Chen X, Zhao LJ, Hu XX, Luo CP, Liang CP, Jiang SW, Liang L, Zheng WC, Guan TP (2019) Impact of livestock terrain utilization patterns on wildlife: A case study of Wanglang National Nature Reserve. Biodiversity Science, 27, 630-637. (in Chinese with English abstract) DOI
	[陈星, 赵联军, 胡茜茜, 罗春平, 梁春平, 蒋仕伟, 梁磊, 郑维超, 官天培 (2019) 基于地形的牲畜空间利用特征及干扰评价——以王朗国家级自然保护区为例. 生物多样性, 27, 630-637.] DOI
[8]	Dufresnes C, Litvinchuk SN (2022) Diversity, distribution and molecular species delimitation in frogs and toads from the Eastern Palaearctic. Zoological Journal of the Linnean Society, 195, 695-760. DOI URL
[9]	Folmer O, Black M, Hoeh W, Lutz R, Vrijenhoek R (1994) DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Molecular Marine Biology and Biotechnology, 3, 294-299. PMID
[10]	Fu JZ, Weadick CJ, Bi K (2007) A phylogeny of the high‐elevation Tibetan megophryid frogs and evidence for the multiple origins of reversed sexual size dimorphism. Journal of Zoology, 273, 315-325. DOI URL
[11]	Gu RH (2005) Analysis and estimation for the landscapes of the nature as the Wanglang National Reserve. Journal of Biology, 22(3), 1-4. (in Chinese with English abstract)
	[顾人和 (2005) 王朗自然保护区的大自然景观类型分析与评价. 生物学杂志, 22(3), 1-4.]
[12]	Gu XD, Liang CP, Dai Q, Li C (2009) A simple and convenient method for measuring and monitoring small terrestrial vertebrates: Drift fence and pitfall traps. Sichuan Journal of Zoology, 28, 273-275, 322. (in Chinese with English abstract)
	[古晓东, 梁春平, 戴强, 李成 (2009) 一种简便实用的小型陆栖脊椎动物监测方法——围栏陷阱法. 四川动物, 28, 273-275, 322.]
[13]	Hebert PDN, Cywinska A, Ball SL, DeWaard JR (2003) Biological identifications through DNA barcodes. Proceedings of the Royal Society B: Biological Sciences, 270, 313-321.
[14]	Hebert PDN, Penton EH, Burns JM, Janzen DH, Hallwachs W (2004a) Ten species in one: DNA barcoding reveals cryptic species in the neotropical skipper butterfly Astraptes fulgerator. Proceedings of the National Academy of Sciences, USA, 101, 14812-14817.
[15]	Hebert PDN, Stoeckle MY, Zemlak TS, Francis CM (2004b) Identification of birds through DNA barcodes. PLoS Biology, 2, e312.
[16]	Hillis DM, Moritz C, Mable BK (1996) Molecular Systematics, 2nd edn. Sinauer Associates Incorporated, Sunderland.
[17]	Hofmann S, Stöck M, Zheng YC, Ficetola FG, Li JT, Scheidt U, Schmidt J (2017) Molecular phylogenies indicate a Paleo-Tibetan origin of Himalayan lazy toads (Scutiger). Scientific Reports, 7, 3308. DOI PMID
[18]	Hou YM, Shi SC, Hu DM, Deng Y, Jiang JP, Xie F, Wang B (2020) A new species of the toothed toad Oreolalax (Anura, Megophryidae) from Sichuan Province, China. ZooKeys, 929, 93-115. DOI URL
[19]	Hu JC, Zhang ZJ, Wei FW (2011) History, current situation and prospects on nature reserves for giant pandas (Ailuropoda melanoleuca) in China. Acta Theriologica Sinica, 31, 10-14. (in Chinese with English abstract)
	[胡锦矗, 张泽钧, 魏辅文 (2011) 中国大熊猫保护区发展历史、现状及前瞻. 兽类学报, 31, 10-14.]
[20]	IUCN (2022) The IUCN Red List of Threatened Species, Version 2022-1. https://www.iucnredlist.org. (accessed on 2022-11-08)
[21]	Ivanova NV, Dewaard JR, Hebert PDN (2006) An inexpensive, automation-friendly protocol for recovering high-quality DNA. Molecular Ecology Notes, 6, 998-1002. DOI URL
[22]	Ji LQ, Jiang JP, Lei FM, Wei FW, Hu YB, Cai B (2022) China checklist of animals. In: Catalogue of Life China: 2022 Annual Checklist. Biodiversity Committee of Chinese Academy of Sciences, Beijing.
[23]	Kang DW, Tan LY, Liu XM, Kang W (2010) Analysis on management status of Wanglang Nature Reserve in Sichuan Province. Journal of Hebei Forestry Science and Technology, (3), 21-23. (in Chinese with English abstract)
	[康东伟, 谭留夷, 刘夏明, 康文 (2010) 四川王朗自然保护区管理状况分析. 河北林业科技, (3), 21-23.]
[24]	Kress WJ, García-Robledo C, Uriarte M, Erickson DL (2015) DNA barcodes for ecology, evolution, and conservation. Trends in Ecology & Evolution, 30, 25-35. DOI URL
[25]	Kumar A, Ghazi MGU, Singh B, Hussain SA, Bhatt D, Gupta SK (2017) Conserve primers for sequencing complete ungulate mitochondrial cytochrome c oxidase I (COI) gene from problematic and decomposed biological samples. Mitochondrial DNA Part B: Resources, 2, 64-66.
[26]	Li AN, Yin GF, Zhang ZJ, Tan JB, Nan X, Ma KP, Guo QH (2018) Space-air-field integrated biodiversity monitoring based on experimental station. Biodiversity Science, 26, 819-827. (in Chinese with English abstract) DOI
	[李爱农, 尹高飞, 张正健, 谭剑波, 南希, 马克平, 郭庆华 (2018) 基于站点的生物多样性星空地一体化遥感监测. 生物多样性, 26, 819-827.] DOI
[27]	Li C, Li J, Liang CP, Zhao LJ, Wang DJ, Jiang JP (2017) The relationship between community structure of ground- dwelling vertebrates and habitat types in the Wanglang Natural Reserve. Acta Ecologica Sinica, 37, 4247-4257. (in Chinese with English abstract)
	[李成, 李娟, 梁春平, 赵联军, 王大军, 江建平 (2017) 王朗自然保护区地栖脊椎动物群落结构和生境类型的关系. 生态学报, 37, 4247-4257.]
[28]	Li GY, Zhang QM (1989) Bird survey in Wanglang Nature Reserve. Sichuan Journal of Zoology, 8(3), 17-20. (in Chinese)
	[李桂垣, 张清茂 (1989) 王朗自然保护区鸟类调查报告. 四川动物, 8(3), 17-20.]
[29]	Li GY, Zhang QM, Wen AX (1993) A description of nests and Passeriformes bird eggs in Sichuan. Sichuan Journal of Zoology, 12(3), 20-26. (in Chinese)
	[李桂垣, 张清茂, 温安祥 (1993) 四川雀形目鸟巢和鸟卵记述. 四川动物, 12(3), 20-26.]
[30]	Li M, Wei FW, Rao G, Fang SG, Feng ZJ (2001) Application of noninvasive sampling in conservation genetics. Acta Theriologica Sinica, 47, 338-342. (in Chinese with English abstract)
	[李明, 魏辅文, 饶刚, 方盛国, 冯祚建 (2001) 非损伤性取样法在保护遗传学研究中的应用. 兽类学报, 47, 338-342.]
[31]	Li S, McShea WJ, Wang DJ, Huang JZ, Shao LK (2012a) A direct comparison of camera-trapping and sign transects for monitoring wildlife in the Wanglang National Nature Reserve, China. Wildlife Society Bulletin, 36, 538-545. DOI URL
[32]	Li S, McShea WJ, Wang DJ, Lu Z, Gu XD (2012b) Gauging the impact of management expertise on the distribution of large mammals across protected areas. Diversity and Distributions, 18, 1166-1176. DOI URL
[33]	Li S, McShea WJ, Wang DJ, Shao LK, Shi XG (2010) The use of infrared-triggered cameras for surveying phasianids in Sichuan Province, China. Ibis, 152, 299-309. DOI URL
[34]	Li S, McShea WJ, Wang DJ, Shen XL, Bu HL, Guan TP, Wang F, Gu XD, Zhang XF, Liao HH (2020) Construction progress of the Camera-trapping Network for the Mountains of Southwest China. Biodiversity Science, 28, 1049-1058. (in Chinese with English abstract) DOI URL
	[李晟, McShea WJ, 王大军, 申小莉, 卜红亮, 官天培, 王放, 古晓东, 张晓峰, 廖灏泓 (2020) 西南山地红外相机监测网络建设进展. 生物多样性, 28, 1049-1058.]
[35]	Liu SY, Ran JH, Lin Q, Liu SC, Chen YP, Jiang SW, Zhao LJ (2001) Diversity of vertebrates in Wanglang Nature Reserve. Journal of Sichuan Forestry Science and Technology, 22(3), 10-14. (in Chinese with English abstract)
	[刘少英, 冉江洪, 林强, 刘世昌, 陈佑平, 蒋仕伟, 赵联军 (2001) 王朗自然保护区脊椎动物多样性. 四川林业科技, 22(3), 10-14.]
[36]	Liu SY, Zhao LJ, Wang X, Liao R (2019) Newly confirmed distribution of Volemys millicens in Wanglang. Journal of Sichuan Forestry Science and Technology, 40(5), 1-4, 10. (in Chinese with English abstract)
	[刘少英, 赵联军, 王新, 廖锐 (2019) 王朗自然保护区发现珍稀兽类四川田鼠. 四川林业科技, 40(5), 1-4, 10.]
[37]	Lohman DJ, Prawiradilaga DM, Meier R (2009) Improved COI barcoding primers for Southeast Asian perching birds (Aves: Passeriformes). Molecular Ecology Resources, 9, 37-40. DOI PMID
[38]	Lü ML, Liu Z, Song Z, Wang YN, Liu XY (2019) Diversity and distribution of culturable Mucoromycota fungi in the Greater Khinggan Mountains, China. Biodiversity Science, 27, 821-832. (in Chinese with English abstract) DOI
	[吕美林, 刘泽, 宋震, 王亚宁, 刘小勇 (2019) 大兴安岭地区可培养毛霉门真菌多样性与分布. 生物多样性, 27, 821-832.] DOI
[39]	Mac Kinnon J, Phillipps K, He FQ (2000) A Field Guide to the Birds of China. Hunan Education Press, Changsha. (in Chinese)
	[约翰·马敬能, 卡伦·菲利普斯, 何芬奇 (2000) 中国鸟类野外手册. 湖南教育出版社, 长沙.]
[40]	Meusnier I, Singer GAC, Landry JF, Hickey DA, Hebert PDN, Hajibabaei M (2008) A universal DNA mini-barcode for biodiversity analysis. BMC Genomics, 9, 214. DOI PMID
[41]	Myers N, Mittermeier RA, Mittermeier CG, da Fonseca GAB, Kent J (2000) Biodiversity hotspots for conservation priorities. Nature, 403, 853-858. DOI
[42]	Palumbi SR, Martin A, Romano S, McMillan WO, Stice L, Grabowski G (2002) The Simple Fool's Guide to PCR, Version 2.0. Department of Zoology and Kewalo Marine Laboratory, Honolulu, Hawaii.
[43]	Parejo-Farnés C, Albaladejo RG, Camacho C, Aparicio A (2018) From species to individuals: Combining barcoding and microsatellite analyses from non-invasive samples in plant ecology studies. Plant Ecology, 219, 1151-1158. DOI
[44]	Parson W, Pegoraro K, Niederstätter H, Föger M, Steinlechner M (2000) Species identification by means of the cytochrome b gene. International Journal of Legal Medicine, 114, 23-28. DOI PMID
[45]	Pyron RA, Wiens JJ (2011) A large-scale phylogeny of Amphibia including over 2800 species, and a revised classification of extant frogs, salamanders, and caecilians. Molecular Phylogenetics and Evolution, 61, 543-583. DOI PMID
[46]	Rao DQ, Wilkinson JA (2008) Phylogenetic relationships of the mustache toads inferred from mtDNA sequences. Molecular Phylogenetics and Evolution, 46, 61-73. DOI URL
[47]	Robins JH, Hingston M, Matisoo-Smith E, Ross HA (2007) Identifying Rattus species using mitochondrial DNA. Molecular Ecology Notes, 7, 717-729. DOI URL
[48]	Rovero F, Tobler M, Sanderson J (2010) Camera trapping for inventorying terrestrial vertebrates. In: Manual on Field Recording Techniques and Protocols for All Taxa Biodiversity inventories and Monitoring (eds Eymann J, Degreef J, Häuser C, Monje JC, Samyn Y, VandenSpiegel D), pp.100-128. The Belgian National Focal Point to the Global Taxonomy initiative, Brussels, Belgium.
[49]	Saitoh T, Sugita N, Someya S, Iwami Y, Kobayashi S, Kamigaichi H, Higuchi A, Asai S, Yamamoto Y, Nishiumi I (2015) DNA barcoding reveals 24 distinct lineages as cryptic bird species candidates in and around the Japanese Archipelago. Molecular Ecology Resources, 15, 177-186. DOI PMID
[50]	Segelbacher G (2002) Noninvasive genetic analysis in birds: Testing reliability of feather samples. Molecular Ecology Notes, 2, 367-369.
[51]	Shan L, Hu YB, Wei FW (2018) Opportunities and challenges of fecal DNA technology in molecular ecology researches. Acta Theriologica Sinica, 38, 235-246. (in Chinese with English abstract) DOI
	[单磊, 胡义波, 魏辅文 (2018) 粪便DNA分析技术在分子生态学研究中的机遇与挑战. 兽类学报, 38, 235-246.]
[52]	Shang XT, Luo CP, Li B, Zheng Y, Zhou ZQ, Zhang L, Li S (2020) Diversity and fauna composition of birds in the Wanglang National Nature Reserve, Sichuan. Sichuan Journal of Zoology, 39, 93-106. (in Chinese with English abstract)
	[尚晓彤, 罗春平, 李斌, 郑勇, 周智强, 张立, 李晟 (2020) 四川王朗国家级自然保护区鸟类多样性与区系组成. 四川动物, 39, 93-106.]
[53]	Shao XN, Lu Q, Liu MZ, Xiong MY, Bu HL, Wang DJ, Liu SY, Zhao JD, Li S, Yao M (2021a) Generalist carnivores can be effective biodiversity samplers of terrestrial vertebrates. Frontiers in Ecology and the Environment, 19, 557-563. DOI URL
[54]	Shao XN, Lu Q, Xiong MY, Bu HL, Shi XY, Wang DJ, Zhao JD, Li S, Yao M (2021b) Prey partitioning and livestock consumption in the world's richest large carnivore assemblage. Current Biology, 31, 4887-4897. DOI URL
[55]	Shao XN, Song DZ, Huang QW, Li S, Yao M (2019) Fast surveys and molecular diet analysis of carnivores based on fecal DNA and metabarcoding. Biodiversity Science, 27, 543-556. (in Chinese with English abstract) DOI
	[邵昕宁, 宋大昭, 黄巧雯, 李晟, 姚蒙 (2019) 基于粪便DNA及宏条形码技术的食肉动物快速调查及食性分析. 生物多样性, 27, 543-556.] DOI
[56]	Simon C, Frati F, Beckenbach A, Crespi B, Liu H, Flook P (1994) Evolution, weighting, and phylogenetic utility of mitochondrial gene sequences and a compilation of conserved polymerase chain reaction primers. Annals of the Entomological Society of America 87, 651-701. DOI URL
[57]	Sourdis J, Nei M (1988) Relative efficiencies of the maximum parsimony and distance-matrix methods in obtaining the correct phylogenetic tree. Molecular Biology and Evolution, 5, 298-311. PMID
[58]	Taberlet P, Coissac E, Pompanon F, Gielly L, Miquel C, Valentini A, Vermat T, Corthier G, Brochmann C, Willerslev E (2007) Power and limitations of the chloroplast trnL (UAA) intron for plant DNA barcoding. Nucleic Acids Research, 35, e14.
[59]	Taberlet P, Waits LP, Luikart G (1999) Noninvasive genetic sampling: Look before you leap. Trends in Ecology & Evolution, 14, 323-327. DOI URL
[60]	Tamura K, Stecher G, Kumar S (2021) MEGA11: Molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution, 38, 3022-3027. DOI PMID
[61]	Tian C, Li JQ, Yang XY, Yu L, Yuan D, Li YX (2018) Preliminary surveys of wild animals using infrared camera in Wanglang National Nature Reserve, Sichuan Province. Biodiversity Science, 26, 620-626. (in Chinese with English abstract) DOI
	[田成, 李俊清, 杨旭煜, 余鳞, 袁丹, 黎运喜 (2018) 野生动物物种多样性的初步调查. 生物多样性, 26, 620-626.] DOI
[62]	Tian C, Liao PC, Dayananda B, Zhang YY, Liu ZX, Li JQ, Yu B, Qing L (2019) Impacts of livestock grazing, topography and vegetation on distribution of wildlife in Wanglang National Nature Reserve, China. Global Ecology and Conservation, 20, e00726.
[63]	Tian J, Zhu SY, Zhang XF, He LW, Gu XD, Guan TP, Li S (2021) The diversity of large- and medium-sized terrestrial mammals and birds in the Giant Panda National Park: A meta-analysis based on camera-trapping data. Biodiversity Science, 29, 1490-1504. (in Chinese with English abstract) DOI
	[田佳, 朱淑怡, 张晓峰, 何礼文, 古晓东, 官天培, 李晟 (2021) 大熊猫国家公园的地栖大中型鸟兽多样性现状: 基于红外相机数据的分析. 生物多样性, 29, 1490-1504.] DOI
[64]	Valentini A, Pompanon F, Taberlet P (2009) DNA barcoding for ecologists. Trends in Ecology & Evolution, 24, 110-117. DOI URL
[65]	Waits LP, Paetkau D (2005) Noninvasive genetic sampling tools for wildlife biologists: A review of applications and recommendations for accurate data collection. Journal of Wildlife Management, 69, 1419-1433. DOI URL
[66]	Wei FW, Rao G, Li M, Fang SG, Feng ZJ (2001) Molecular scatology and its application—Reliability, limitation and prospect. Acta Theriologica Sinica, 21, 143-152, 160. (in Chinese with English abstract)
	[魏辅文, 饶刚, 李明, 方盛国, 冯祚建 (2001) 分子粪便学及其应用——可靠性、局限性和展望. 兽类学报, 21, 143-152, 160.]
[67]	Wei XR (1988) Supplement of small mammals in Wanglang Nature Reserve of Pingwu County. Sichuan Journal of Zoology, 7, 42. (in Chinese)
	[魏孝荣 (1988) 平武县王朗自然保护区小型兽补遗. 四川动物, 7, 42.
[68]	Wei XR (1991) A list of fleas and mites in vitro from small mammals in Wanglang Nature Reserve of Pingwu, Sichuan Province. Chinese Journal of Vector Biology and Control, 2, 126. (in Chinese)
	[魏孝荣 (1991) 四川省平武王朗自然保护区小型兽体外蚤、螨名录. 中国媒介生物学及控制杂志, 2, 126.]
[69]	Wu H, Xu XH, Feng XJ, Mi XC, Su YJ, Xiao ZS, Zhu CD, Cao L, Gao X, Song CY, Guo LD, Wu DH, Jiang JP, Shen H, Ma KP (2022) Progress and prospect of China biodiversity monitoring from a global perspective. Biodiversity Science, 30, 22434. (in Chinese with English abstract) DOI
	[吴慧, 徐学红, 冯晓娟, 米湘成, 苏艳军, 肖治术, 朱朝东, 曹垒, 高欣, 宋创业, 郭良栋, 吴东辉, 江建平, 沈浩, 马克平 (2022) 全球视角下的中国生物多样性监测进展与展望. 生物多样性, 30, 22434.] DOI
[70]	Xiong MY, Shao XN, Long Y, Bu HL, Zhang D, Wang DJ, Li S, Wang RJ, Yao M (2016) Molecular analysis of vertebrates and plants in scats of leopard cats (Prionailurus bengalensis) in southwest China. Journal of Mammalogy, 97, 1054-1064. DOI URL
[71]	Ye CY, Fei L (2007) A new species of Megophryidae—Scutiger (Scutiger) wanglangensis from Sichuan, China (Amphibia, Anura). Herpetologica Sinica, 11, 33-37. (in Chinese with English abstract)
	[叶昌媛, 费梁 (2007) 四川角蟾科(Megophryidae)一新种——王朗齿突蟾(两栖纲: 无尾目). 两栖爬行动物学研究, 11, 33-37.]
[72]	Yu KX, Hu QQ, Yao YA, Zhang SS, Xu AC, Ge J, Guan F (2022) Research progress on wildlife identification technology. Bulletin of Science and Technology, 38(2), 1-10, 18. (in Chinese with English abstract)
	[俞可心, 胡情情, 姚逸安, 章书声, 徐爱春, 葛建, 管峰 (2022) 野生动物鉴定技术研究进展. 科技通报, 38(2), 1-10, 18.]
[73]	Zhan XJ (2006) Using Noninvasive Genetic Sampling to Estimate the Population Size and Study Dispersal of Giant Pandas. PhD dissertation, Institute of Zoology, Chinese Academy of Sciences, Beijing. (in Chinese with English abstract)
	[詹祥江 (2006) 利用非损伤性遗传取样研究大熊猫的种群数量和扩散模式. 博士学位论文, 中国科学院动物研究所, 北京.]
[74]	Zhang BW, Wei FW, Li M, Lü XP (2004) A simple protocol for DNA extraction from faeces of the giant panda and lesser panda. Acta Theriologica Sinica, 50, 452-458. (in Chinese with English abstract)
	[张保卫, 魏辅文, 李明, 吕晓平 (2004) 大熊猫和小熊猫粪便DNA提取的简易方法. 兽类学报, 50, 452-458.]
[75]	Zhang GX, Wang ZP, Zhong ZM, Feng YW (1991) A survey of small mammals in Wanglang Nature Reserve. Sichuan Journal of Zoology, 10(2), 41. (in Chinese)
	[张国修, 王再平, 钟肇敏, 冯云武 (1991) 王朗自然保护区小型兽类的调查. 四川动物, 10(2), 41.]
[76]	Zhang WY, Shu GC, Li YL, Xiong S, Liang CP, Li C (2018) Daytime driving decreases amphibian roadkill. PeerJ, 6, e5385.
[77]	Zhang YP, Wang XX, Ryder OA, Li HP, Zhang HM, Yong YG, Wang PY (2002) Genetic diversity and conservation of endangered animal species. Pure and Applied Chemistry, 74, 575-584. DOI URL
[78]	Zhao LJ, Li XR, Sun ZY, Jiang SW, Chen YP, Liu SY (2016) A study of diversities of small mammals in Wanglang National Nature Reserve. Journal of Sichuan Forestry Science and Technology, 37(3), 66-68, 14. (in Chinese with English abstract)
	[赵联军, 李小蓉, 孙志宇, 蒋仕伟, 陈幼平, 刘少英 (2016) 王朗国家级自然保护区小型兽类多样性研究. 四川林业科技, 37(3), 66-68, 14.]
[79]	Zheng YC, Mo BH, Liu ZJ, Zeng XM (2004) Phylogenetic relationships of megophryid genera (Anura: Megophryidae) based on partial sequences of mitochondrial 16S rRNA gene. Zoological Research, 25, 205-213. (in Chinese with English abstract)
	[郑渝池, 莫邦辉, 刘志君, 曾晓茂 (2004) 基于16S rRNA序列的角蟾科部分属间系统关系. 动物学研究, 25, 205-213.]

类别 Category	目标片段所在基因(引物名称) Target gene (primers' names)	上游引物序列5'-3' Forward primer sequence 5'-3'	下游引物序列5'-3' Reverse primer sequence 5'-3'	退火温度 Annealing temperature (℃)	循环次数 Cycles	参考文献 Reference
兽类 Mammals	COI (COIung)	GTACCGCTAATAATTGGTGCTCC	GGGTGGCCAAAGAATCAGAACAAGTG	56	35	Kumar et al, 2017
	16S (16SF/R)	GAGAAGACCCTATGGAGC	ATAGAAACCGACCTGGAT	50	30	Xiong et al, 2016
	COI (BatL/R6036R)	CCTACTCRGCCATTTTACCTATG	ATCTCTGGGTGTCCAAAGAATCA	55	35	Robins et al, 2007
	ATP6 (ATP6-DF3/ATP6-DR2)	AACGAAAATCTATTCGCCTCT	TGGATGGACAGTATTTGTTTTGAT	60/50	10/30	Chaves et al, 2012
	D-loop (BEDL225/H470)	ATGTACATACTGTGCTTGGC	GTCATTAGTCCATCGAGATG	53	38	Zhang et al, 2002
鸟类 Avians	COI (COI-K_Bird_F/R)	CCCCAGACATAGCATTYCC	TTGTGATAGTGGTGGGGTTTTAT	46/53	5/45	Parejo-Farnés et al, 2018
	Cytb (Cytb-L14816/H15173)	CCATCCAACATCTCAGCATGATGAAA	CCCCTCAGAATGATATTTGTCCTCA	50	30	Parson et al, 2000
	COI (L6697Bird_F/H7390Thrush_R)	TCAACYAACCACAAAGAYATCGGYAC	ACGTGGGARATRATTCCAAATCCTG	48/51	5/30	Saitoh et al, 2015
	COI (PasserF/R)	CCAACCACAAAGACATCGGAACC	GTAAACTTCTGGGTGACCAAAGAATC	58	35	Lohman et al, 2009
两栖类 Amphibians	COI (LCO1490/HCO2198)	GGTCAACAAATCATAAAGATATTGG	TAAACTTCAGGGTGACCATCA	50	35	Folmer et al, 1994
	COI (LepF/R)	ATTCAACCAATCATAAAGATATTGG	TAAACTTCTGGATGTCCATCA	46	35	Hebert et al, 2004a
	16S (P7/8)	CGCCTGTTTACCCAT	CCGGTCTGAACTCAGATCACGT	55	36	Simon et al, 1994
	16S (Sar/Sbr)	CGCCTGTTTATCCAT	CCGGTCTGAACTCAGATCACGT	48	40	Palumbi et al, 2002
	12S-16S (1602L/2571H)	GTATACCGGAAGGTGTACTTGGAACAG	TACCTTCGCACGGTCAGAATACCGC	50	35	Fu et al, 2007
	Cytb (FrogGlu-f/Thr-r)	TGATCTGCCACCGTTG	CTCCATTCTTCGRCTTACAAG	46	40	Hillis et al, 1996
	RAG1 (RAG1-F/R)	AGCTGCAGYCARTACCAYAARATGTA	GCAAAGTTTCCGTTCATTCTCAT	50	35	Fu et al, 2007

类别 Category	目标片段所在基因(引物名称) Target gene (primers' names)	上游引物序列5'-3' Forward primer sequence 5'-3'	下游引物序列5'-3' Reverse primer sequence 5'-3'	退火温度 Annealing temperature (℃)	循环次数 Cycles	参考文献 Reference
兽类 Mammals	COI (COIung)	GTACCGCTAATAATTGGTGCTCC	GGGTGGCCAAAGAATCAGAACAAGTG	56	35	Kumar et al, 2017
	16S (16SF/R)	GAGAAGACCCTATGGAGC	ATAGAAACCGACCTGGAT	50	30	Xiong et al, 2016
	COI (BatL/R6036R)	CCTACTCRGCCATTTTACCTATG	ATCTCTGGGTGTCCAAAGAATCA	55	35	Robins et al, 2007
	ATP6 (ATP6-DF3/ATP6-DR2)	AACGAAAATCTATTCGCCTCT	TGGATGGACAGTATTTGTTTTGAT	60/50	10/30	Chaves et al, 2012
	D-loop (BEDL225/H470)	ATGTACATACTGTGCTTGGC	GTCATTAGTCCATCGAGATG	53	38	Zhang et al, 2002
鸟类 Avians	COI (COI-K_Bird_F/R)	CCCCAGACATAGCATTYCC	TTGTGATAGTGGTGGGGTTTTAT	46/53	5/45	Parejo-Farnés et al, 2018
	Cytb (Cytb-L14816/H15173)	CCATCCAACATCTCAGCATGATGAAA	CCCCTCAGAATGATATTTGTCCTCA	50	30	Parson et al, 2000
	COI (L6697Bird_F/H7390Thrush_R)	TCAACYAACCACAAAGAYATCGGYAC	ACGTGGGARATRATTCCAAATCCTG	48/51	5/30	Saitoh et al, 2015
	COI (PasserF/R)	CCAACCACAAAGACATCGGAACC	GTAAACTTCTGGGTGACCAAAGAATC	58	35	Lohman et al, 2009
两栖类 Amphibians	COI (LCO1490/HCO2198)	GGTCAACAAATCATAAAGATATTGG	TAAACTTCAGGGTGACCATCA	50	35	Folmer et al, 1994
	COI (LepF/R)	ATTCAACCAATCATAAAGATATTGG	TAAACTTCTGGATGTCCATCA	46	35	Hebert et al, 2004a
	16S (P7/8)	CGCCTGTTTACCCAT	CCGGTCTGAACTCAGATCACGT	55	36	Simon et al, 1994
	16S (Sar/Sbr)	CGCCTGTTTATCCAT	CCGGTCTGAACTCAGATCACGT	48	40	Palumbi et al, 2002
	12S-16S (1602L/2571H)	GTATACCGGAAGGTGTACTTGGAACAG	TACCTTCGCACGGTCAGAATACCGC	50	35	Fu et al, 2007
	Cytb (FrogGlu-f/Thr-r)	TGATCTGCCACCGTTG	CTCCATTCTTCGRCTTACAAG	46	40	Hillis et al, 1996
	RAG1 (RAG1-F/R)	AGCTGCAGYCARTACCAYAARATGTA	GCAAAGTTTCCGTTCATTCTCAT	50	35	Fu et al, 2007

类别 Category	数量 No. of samples	样品类型 Sample type	数量 No. of samples
鸟类 Avians	156	粪便 Feces	123
兽类 Mammals	137	羽毛(毛发) Feathers (Pelage)	60
两栖类 Amphibians	21	组织 Tissues	37
		标本组织 Tissues from specimens	90
		卵壳 Eggshells	4
总计 Total	314	总计 Total	314

类别 Category	数量 No. of samples	样品类型 Sample type	数量 No. of samples
鸟类 Avians	156	粪便 Feces	123
兽类 Mammals	137	羽毛(毛发) Feathers (Pelage)	60
两栖类 Amphibians	21	组织 Tissues	37
		标本组织 Tissues from specimens	90
		卵壳 Eggshells	4
总计 Total	314	总计 Total	314

物种类型(样品数量/个) Category (No. of samples)	成功扩增样品数(扩增成功率) No. of successfully amplified samples (Success rate)	通用引物鉴定物种数(识别率) No. of species identified by universal primers (Identification rate)	特异性引物鉴定物种数(识别率) No. of species identified by specific primers (Identification rate)
鸟类 Avians (156)	128 (82.1%)	41 (89.1%)	5 (10.9%)
兽类 Mammals (137)	111 (81.0%)	15 (55.6%)	12 (44.4%)
两栖类 Amphibians (21)	21 (100%)	3 (75.0%)	-

基于DNA条形码技术构建王朗国家级自然保护区陆生脊椎动物遗传资源数据库及物种鉴定

The establishment of terrestrial vertebrate genetic resource bank and species identification based on DNA barcoding in Wanglang National Nature Reserve

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物种类别 Category	样品数 No. of samples	物种预判样品数 No. of morphologically identified samples	预判正确样品数 No. of correctly identified samples	预判正确率 Accuracy of morphological identification rate
鸟类 Avians	109	57	30	52.6%
兽类 Mammals	99	38	18	47.4%
两栖类 Amphibians	13	13	9	69.2%
合计 Total	221	108	57	52.8%

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