Identifying homomorphic sex chromosomes from wild‐caught adults with limited genomic resources

Abstract

We demonstrate a genotyping-by-sequencing approach to identify homomorphic sex chromosomes and their homolog in a distantly related reference genome, based on noninvasive sampling of wild-caught individuals, in the moor frog Rana arvalis. Double-digest RADseq libraries were generated using buccal swabs from 30 males and 21 females from the same population. Search for sex-limited markers from the unfiltered data set (411 446 RAD tags) was more successful than searches from a filtered data set (33 073 RAD tags) for markers showing sex differences in heterozygosity or in allele frequencies. Altogether, we obtained 292 putatively sex-linked RAD loci, 98% of which point to male heterogamety. We could map 15 of them to the Xenopus tropicalis genome, all but one on chromosome pair 1, which seems regularly co-opted for sex determination among amphibians. The most efficient mapping strategy was a three-step hierarchical approach, where R. arvalis reads were first mapped to a low-coverage genome of Rana temporaria (17 My divergence), then the R. temporaria scaffolds to the Nanorana parkeri genome (90 My divergence), and finally the N. parkeri scaffolds to the X. tropicalis genome (210 My). We validated our conclusions with PCR primers amplifying part of Dmrt1, a candidate sex determination gene mapping to chromosome 1: a sex-diagnostic allele was present in all 30 males but in none of the 21 females. Our approach is likely to be productive in many situations where biological samples and/or genomic resources are limited.

Are glacial refugia hotspots of speciation and cytonuclear discordances? Answers from the genomic phylogeography of Spanish common frogs

“…The library was sequenced on three Illumina HiSeq 2500 lanes (single read 125) and raw sequences were demultiplexed with Stacks v1.48 (Catchen, Hohenlohe, Bassham, Amores, & Cresko, ). We used the denovo.pl pipeline with default parameters ( –M 2, –m 3 and –n 2) to catalogue the tags sequenced in each sample, including additional RAD sequence reads from five individuals of R. arvalis (a close relative of R. temporaria ) obtained with the same protocol (Brelsford, Lavanchy, Sermier, Rausch, & Perrin, ), to be used as outgroups (Rarv49, Rarv70, Rarv92, Rarv66, Rarv81). For population genomic analyses across northern Spain, single nucleotide polymorphisms (SNPs) were called from RAD tags (118 bp sequences) sequenced in all populations (– p 33) and in all samples of each (– r 1), while discarding those bearing rare variants ( –min_maf 0.05) and those that were predominantly heterozygous ( –max_het_obs 0.75), which can represent overmerged paralogues.…”

Sex chromosome turnovers and genetic drift: a simulation study

“…Sex chromosome turnovers have also occurred multiple times in Ranid frogs, but only one has resulted in a change in the heterogametic sex (Sumida & Nishioka, 2000;Miura, 2008). Furthermore, the rate of turnovers that preserve the heterogamety pattern (XY to XY and ZW to ZW turnovers) is likely underappreciated in many taxa because of technical limitations (Brelsford et al, 2017). Some of the currently used methods to detect sex determination rely solely on the identification of sex-linked markers (which allow to distinguish between male and female heterogamety) but does not single out the sex chromosome pair, preventing the identification of turnovers within groups homogeneous for heterogamety (Gamble et al, 2015;Lambert et al, 2016).…”

Sex is determined by XY chromosomes across the radiation of dioeciousNepenthespitcher plants

“…These strategies can potentially allow sex-linked regions to be discovered by cheaper reduced-representation sequencing (RRS) methods such as RAD-seq (Baird et al 2008;Elshire et al 2011;Peterson et al 2012), although the gained information will remain incomplete, because typically only a few percent of a genome is covered. Nevertheless, the discovery of sex-linked markers by RRS has been successful in organisms such as Crustaceans (Carmichael et al 2013), Anolis lizards (Gamble and Zarkower 2014), geckos (Gamble et al 2015a), and frogs (Brelsford et al 2017;Jeffries et al 2018).…”

“…A major problem faced by approaches that use population polymorphism to infer sex-linkage, and sex specificity in particular, is error in the measurement of locus presence and absence (Text S4). Presence-absence error has long been recognized as a problem in fragment length genotyping methods, but it is exacerbated in RRS data, in which missing loci occur in a highly stochastic manner (Mastretta-Yanes et al 2015;Bresadola et al 2019), and can make sex-specific sequences appear in both sexes (Bewick et al 2013;Gamble and Zarkower 2014;Heikrujam et al 2015;Brelsford et al 2017), and probably represent false positive results. One suggested solution to reduce the number of false positives is to compare increasing numbers of males and females (Gamble and Zarkower 2014;Gamble et al 2015b).…”