体表擦拭取样在两栖动物保护遗传学研究中的应用
The Application of Extracting DNA from Skin Swabbing in Amphibians

中文摘要：相比于其他取样方法，非损伤性取样对动物的伤害可以降到最低，因而在动物保护研究中有着重要的价值。在两栖类中通过体表擦拭进行非损伤性取样的方法已有报道，但是相关研究仅局限于零星物种，且均是在实验室环境下完成取样，并不适用于野外就地取样。本研究对四川申果庄省级大熊猫自然保护区内有尾两栖类及无尾两栖类6科10属11种进行了就地擦拭取样。线粒体基因序列分析表明，体表擦拭取样法获得的DNA能满足常规的分子生物学实验要求，可用于水生、陆栖等各种两栖动物类群，可在两栖类保护遗传学研究中广泛应用。
英文摘要：Compared to other sampling methods, non-invasive sampling can minimize the damage to animals, so it has important value for animal protection. There is only a few reports on skin swabbing in a limited number of amphibian species, and the sampling methods are all performed in laboratory environment and are not suitable for application in the wild fields. In this study, Shenguozhuang Provincial Giant Panda Nature Reserve in Sichuan was selected as the research area. The skin swabbing method was used on wild populations of 11 amphibian species to test DNA yields and quality. Finally, the DNA was successfully obtained and multiple of genetic markers were amplified. Based on PCR performance, DNA yields and sampling considerations, the method of skin swab sampling was confirmed to work well to obtain DNA without harming the amphibian. Therefore, skin swabbing is a reliable method for obtaining genetic data from endangered amphibian without jeopardizing their survival. 2018,37(4): 373-380 收稿日期：2017-09-23 DOI：10.11984/j.issn.1000-7083.20170293 分类号：Q959.5 基金项目：国家自然科学基金项目（NSFC31572241）；"环境保护部生物多样性专项"支持项目 作者简介：艾永斌,男,林业工程师,研究方向为野生动物与自然保护区管理,E-mail:352278596@qq.com *通讯作者:曾晓茂,E-mail:zengxm@cib.ac.cn 参考文献： 费梁, 胡淑琴, 叶娼媛, 等. 2009. 中国动物志 两栖纲[M]. 北京:科学出版社. 李霞, 陈新诺, 王蕾, 等. 2015. 山羊粪便总DNA提取方法的比较[J]. 四川畜牧兽医, 42(1):38-40. 邵劲松, 浦牧野, 顾祝军. 2013. 3种土壤微生物基因组DNA提取方法的比较[J]. 南京晓庄学院学报, 29(3):64-67. 吴军, 高逖, 徐海根, 等. 2013. 两栖动物监测方法和国外监测计划研究[J]. 生态与农村环境学报, 29(6):784-788. Broquet T, Berset-Braendli L, Emaresi G, et al. 2007. Buccal swabs allow efficient and reliable microsatellite genotyping in amphibians[J]. Conservation Genetics, 8(2):509-511. Che J, Chen HM, Yang JX, et al. 2012. Universal COⅠ primers for DNA barcoding amphibians[J]. Molecular Ecology Resources, 12(2):247-258. Frost DR. 2017. Amphibian species of the world:an online reference version 5.6[EB/OL]. (2013/01/09). http://research.amnh.org/herpetology/amphibia/index.html. Guo W, Ao M, Li W, et al. 2013. Skin secretion and shedding is a good source for non-destructive genetic sampling in the Chinese giant salamander ( Andrias davidianus)[J]. Zeitschrift für Naturforschung C, Journal of Biosciences, 68(3-4):164-108. Ivanova NV, Dewaard JR, Hebert PDN. 2006. An inexpensive, automation-friendly protocol for recovering high-quality DNA[J]. Molecular Ecology Resources,