Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
Mol Ecol Resour. 2012 Nov;12(6):1105-13. doi: 10.1111/1755-0998.12000. Epub 2012 Sep 1.
Studies of population genetics increasingly use next-generation DNA sequencing to identify microsatellite loci in nonmodel organisms. There are, however, relatively few studies that validate the feasibility of transitioning from marker development to experimental application across populations and species. North American coralsnakes of the Micrurus fulvius species complex occur in the United States and Mexico, and little is known about their population structure and phylogenetic relationships. This absence of information and population genetics markers is particularly concerning because they are highly venomous and have important implications on human health. To alleviate this problem in coralsnakes, we investigated the feasibility of using 454 shotgun sequences for microsatellite marker development. First, a genomic shotgun library from a single individual was sequenced (approximately 7.74 megabases; 26,831 reads) to identify potentially amplifiable microsatellite loci (PALs). We then hierarchically sampled 76 individuals from throughout the geographic distribution of the species complex and examined whether PALs were amplifiable and polymorphic. Approximately half of the loci tested were readily amplifiable from all individuals, and 80% of the loci tested for variation were variable and thus informative as population genetic markers. To evaluate the repetitive landscape characteristics across multiple snakes, we also compared microsatellite content between the coralsnake and two other previously sampled snakes, the venomous copperhead (Agkistrodon contortrix) and Burmese python (Python molurus).
越来越多的群体遗传学研究利用下一代 DNA 测序来鉴定非模式生物中的微卫星基因座。然而,相对较少的研究验证了从标记开发过渡到跨群体和物种的实验应用的可行性。北美珊瑚蛇属于 Micrurus fulvius 物种复合体,分布于美国和墨西哥,人们对其种群结构和系统发育关系知之甚少。由于这些蛇类具有高度毒性,对人类健康有重要影响,而有关其种群遗传标记的信息却相对较少,这令人尤为担忧。为了解决珊瑚蛇的这一问题,我们研究了使用 454 焦磷酸测序法开发微卫星标记的可行性。首先,对单个个体的基因组进行鸟枪法测序(约 774 兆碱基;26831 条reads),以鉴定潜在可扩增的微卫星基因座(PALs)。然后,我们在物种复合体的整个地理分布范围内对 76 个个体进行分层采样,并检验 PALs 是否可扩增和多态性。经测试的大约一半的微卫星基因座可从所有个体中轻易扩增,且有 80%用于检测变异的微卫星基因座具有变异性,因此可用作种群遗传标记。为了评估多个蛇种的重复序列特征,我们还比较了珊瑚蛇与另外两种先前采样的蛇种(毒性铜斑蛇 Agkistrodon contortrix 和缅甸蟒 Python molurus)之间的微卫星内容。