Abstract
Multiple nuclear markers provide genetic polymorphism data for molecular systematics and population genetic studies. They are especially required for the coalescent-based analyses that can be used to accurately estimate species trees and infer population demographic histories. However, in avian evolutionary studies, these powerful coalescent-based methods are hindered by the lack of a sufficient number of markers. In this study, we designed PCR primers to amplify 136 nuclear protein-coding loci (NPCLs) by scanning the published Red Junglefowl (Gallus gallus) and Zebra Finch (Taeniopygia guttata) genomes. To test their utility, we amplified these loci in 41 bird species representing 23 Aves orders. The sixty-three best-performing NPCLs, based on high PCR success rates, were selected which had various mutation rates and were evenly distributed across 17 avian autosomal chromosomes and the Z chromosome. To test phylogenetic resolving power of these markers, we conducted a Neoavian phylogenies analysis using 63 concatenated NPCL markers derived from 48 whole genomes of birds. The resulting phylogenetic topology, to a large extent, is congruence with results resolved by previous whole genome data. To test the level of intraspecific polymorphism in these makers, we examined the genetic diversity in four populations of the Kentish Plover (Charadrius alexandrinus) at 17 of NPCL markers chosen at random. Our results showed that these NPCL markers exhibited a level of polymorphism comparable with mitochondrial loci. Therefore, this set of pan-avian nuclear protein-coding loci has great potential to facilitate studies in avian phylogenetics and population genetics.
Highlights
The generation sequencing technologies have produced sequences data in the unprecedented quantity with relative low cost[1], traditional Sanger sequencing still has its niche in molecular evolutionary studies: pilot or small scale phylogenetic studies using PCR-based approach are cost-effective and nearly available for every laboratory, beneficial to design sampling strategy and built an analysis scheme
We aimed to develop a set of avian universal NPCL markers that can be widely utilized in avian phylogenetic and population genetic studies
More than one primer pairs were conducted for each NPCL marker candidate, and we chose the pair of PCR markers with the highest score denoting the level of conservatism between Zebra Finch and Red Junglefowl genomes
Summary
The generation sequencing technologies have produced sequences data in the unprecedented quantity with relative low cost[1], traditional Sanger sequencing still has its niche in molecular evolutionary studies: pilot or small scale phylogenetic studies using PCR-based approach are cost-effective and nearly available for every laboratory, beneficial to design sampling strategy and built an analysis scheme. A set of universal nuclear markers could provide an efficient way to ease this time consuming process It should greatly facilitate the use of coalescent-based analyses to answer phylogenetic and population genetic questions[5]. The development of a set of universal NPCL markers for birds should significantly reduce the time required for future research as well as its cost, and facilitate the application of coalescent-based methods in avian evolutionary studies. We aimed to develop a set of avian universal NPCL markers that can be widely utilized in avian phylogenetic and population genetic studies. To test the resolving power of these markers, we further constructed a phylogenetic tree and estimated mutation rates by extracting universal NPCLs from 48 published avian genomes[41]. Samples from four populations of the Kentish Plover (Charadrius alexandrinus) were amplified to estimate the intra-specific polymorphic level of these universal NPCLs
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