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Gene gains and losses in the Drosophila Y. Gene location ( Y-linkage vs. autosomal/X-linkage) was determined by PCR. Direction of the movements (gene gains, red arrows; gene losses, blue arrows) was inferred by synteny and parsimony (Supplemental Figs. S1–S4). For the six ancestral genes (dashed arrows), there is no close outgroup for inferring the direction (gain versus loss) (Koerich et al. 2008). Genes were labeled with the names of the D. melanogaster ( JYalpha, CG11719, CG2964) or D. virilis (GJ19835) orthologs. Data for genes JYalpha (abridged as ''JYa'' ), GJ19835, CG11719, and CG2964 came from the present study, and the remaining genes from Koerich et al. (2008).
Source publication
Notwithstanding their biological importance Y chromosomes remain poorly known in most species. A major obstacle to their study is the identification of Y chromosome sequences: due to its high content of repetitive DNA, in most genome projects the Y chromosome sequence is fragmented into a large number of small, unmapped scaffolds. Identification of...
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... the Y-chromosome ( Carvalho et al. 2009). In order to determine when they were acquired by the Y-chromosome, we investigated other Drosophila species using PCR and synteny analysis as described in Koerich et al. (2008). Three genes (GJ19835, GJ19633, and GJ11126) moved to the Y-chromosome after the split between D. melanogaster and D. virilis ( Fig. 4; Supplemental Figs. S1-S4; Supplemental Table S2). In contrast, we found that GJ18574/JYalpha belong to the ancestral Drosophila Y-chromosome; its autosomal location in D. melanogaster resulted from a Y-to-autosome movement in the ancestor of the melanogaster subgroup of ...
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... well-supported in mammals and other groups (e.g., Skaletsky et al. 2003), the evidence in Drosophila does not support it (Carvalho et al. 2009). For example, a previous study with ten Drosophila species ( Koerich et al. 2008) estimated that the rate of gene gain is 10.7-fold higher than the rate of gene loss (95% confidence in- terval: 2.2-51.3 (Fig. 4). These values imply a gene gain/gene loss ratio of 11.0 (P = 0.022, Poisson regression) ( Table 1; Supplemental Methods). Similar results were obtained by applying the method described in Koerich et al. (2008) to the combined D. virilis and D. melanogaster data or to each species separately (Table 1) and by approximate Bayesian ...
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... the difference between Y-linked and not Y-linked small scaffolds; this problem is much ameliorated by k-mer validation (Supplemental Fig. S16). Contaminant scaffolds are spuriously detected as Y-linked if control male reads are not used (Supplemental Fig. S9). Assemblers differ in their performance, sometimes dramatically (e.g., Supplemental Fig. S14), but unfortunately it is difficult to obtain a reliable measure of assembly quality without a reference genome ( Salzberg et al. 2012). Thus, as in all analyses of unfinished genomes, the YGS perfor- mance may be affected by assembly errors that are not easy to pre- dict in ...
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... coverage introduces significant binomial sampling error and hence reduces resolution for small scaffolds. It is difficult to provide a more precise guidance on the minimum female se- quencing coverage required to accurately detect Y-linked scaffolds because it depends on many factors such as genome properties (e.g., amount of segmental duplications), assembly quality (e.g., fragmentation), and the error rates in the genome and female short reads (e.g., Supplemental Fig. S14). ...
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... primarily designed to detect Y-linked (or W-linked) scaffolds, YGS seems to be a useful tool for detection of contami- nant (Supplemental Fig. S10A) and low-quality scaffolds (Supple- mental Fig. S14), and for assembly comparisons/detection of new sequences in the presence of large amounts of repetitive DNA (Supplemental Fig. ...
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... doing the removal of sequencing errors (see previous section), we noticed that the CAF1.2 assembly has a very large number of low quality, small scaffolds. Namely, the CAF1.2 assembly has 13,530 scaffolds, and only 2261 have >80% valid single-copy k-mers ( Supplemental Fig. S14A); among these 2261, only 1056 have >50 valid single-copy k-mers. As detailed in the previous section, we did this comparison using Sanger traces filtered at a phred score of 20 (i.e., 1% error), so ;11,300 scaffolds (i.e., 13,530 minus 2261) contain a substantial amount of low quality sequence. ...
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... on exactly the same Sanger traces. We examined two of them, ''CA'' (Celera Assembler) and ''AR'' (Arachne), using the same procedures we did with the CAF1.2 assembly. We found that the AR assembly was similar to CAF1.2, containing a large number of small, low quality scaffolds (not shown), which are absent from the CA assembly (Supple- mental Fig. S14, panels C,D). Unless otherwise noted, we used the CA assembly in our analyses to avoid the use of filtered data. It is similar to the filtered CAF1.2 assembly both in number of scaf- folds (1186) and in size (165 Mbp; all reported assembly sizes ex- clude gaps). The main results (namely, the identification of four new Y-linked genes and 11 ...
Citations
... To identify candidate sex chromosome regions, we employed the Y chromosome genome scan (YGS) method [68], which was previously used to identify Drosophila melanogaster sex chromosome contigs. Briefly, reads from the same sex were pooled, and 15-mers were extracted with jellyfish count v2.2.4 or v2.2.10 [69]. ...
Background
In gonochoristic animals, the sex determination pathway induces different morphological and behavioral features that can be observed between sexes, a condition known as sexual dimorphism. While many components of this sex differentiation cascade show high levels of diversity, factors such as the Doublesex-Mab-3-Related Transcription factor (DMRT) are widely conserved across animal taxa. Species of the phylum Tardigrada exhibit remarkable diversity in morphology and behavior between sexes, suggesting a pathway regulating this dimorphism. Despite the wealth of genomic and zoological knowledge accumulated in recent studies, the sexual differences in tardigrades genomes have not been identified. In the present study, we focused on the gonochoristic species Paramacrobiotus metropolitanus and employed omics analyses to unravel the molecular basis of sexual dimorphism.
Results
Transcriptome analysis between sex-identified specimens revealed numerous differentially expressed genes, of which approximately 2,000 male-biased genes were focused on 29 non-male-specific genomic loci. From these regions, we identified two Macrobiotidae family specific DMRT paralogs, which were significantly upregulated in males and lacked sex specific splicing variants. Furthermore, phylogenetic analysis indicated all tardigrade genomes lack the doublesex ortholog, suggesting doublesex emerged after the divergence of Tardigrada. In contrast to sex-specific expression, no evidence of genomic differences between the sexes was found. We also identified several anhydrobiosis genes that exhibit sex-biased expression, suggesting a possible mechanism for protection of sex-specific tissues against extreme stress.
Conclusions
This study provides a comprehensive analysis for analyzing the genetic differences between sexes in tardigrades. The existence of male-biased, but not male-specific, genomic loci and identification of the family specific male-biased DMRT subfamily provides the foundation for understanding the sex determination cascade. In addition, sex-biased expression of several tardigrade-specific genes which are involved their stress tolerance suggests a potential role in protecting sex-specific tissue and gametes.
... The simplest way to distinguish sex-linked loci from autosomal ones is to identify those found in reads that mapped to the sex chromosomes of the reference genome. However, this is not possible when (i) a reference genome is not available-as is the case for most wildlife species-and de novo genotyping is required, (ii) there is little conserved synteny between the studied genome and the reference, or (iii) the W/Y chromosome of the reference genome is fragmented into numerous unmapped scaffolds, as is common in many genome projects (Carvalho & Clark, 2013). ...
Identifying sex-linked markers in genomic datasets is important because their presence in supposedly neutral autosomal datasets can result in incorrect estimates of genetic diversity, population structure and parentage. However, detecting sex-linked loci can be challenging, and available scripts neglect some categories of sex-linked variation. Here, we present new R functions to (1) identify and separate sex-linked loci in ZW and XY sex determination systems and (2) infer the genetic sex of individuals based on these loci. We tested these functions on genomic data for two bird and one mammal species and compared the biological inferences made before and after removing sex-linked loci using our function. We found that our function identified autosomal loci with ≥98.8% accuracy and sex-linked loci with an average accuracy of 87.8%. We showed that standard filters, such as low read depth and call rate, failed to remove up to 54.7% of sex-linked loci. This led to (i) overestimation of population FIS by up to 24%, and the number of private alleles by up to 8%; (ii) wrongly inferring significant sex differences in heterozygosity; (iii) obscuring genetic population structure and (iv) inferring ~11% fewer correct parentages. We discuss how failure to remove sex-linked markers can lead to incorrect biological inferences (e.g. sex-biased dispersal and cryptic population structure) and misleading management recommendations. For reduced-representation datasets with at least 15 known-sex individuals of each sex, our functions offer convenient resources to remove sex-linked loci and to sex the remaining individuals (freely available at https://github.com/drobledoruiz/conservation_genomics).
... It is well known that the Y chromosomes of Drosophila and mammals, and the W chromosomes 18 of birds carry only small fractions of the genes carried by the homologous X or Z chromosomes, 19 and this "genetic degeneration" is associated with loss of recombination between the sex 20 chromosome pair. However, it is still not known how much evolutionary time is needed to 21 reach such nearly complete degeneration. ...
... picta, M. parae and M. bifurca) have 16 highly degenerated Ys , and, as reviewed in Fong, et al. (2023), who use the genus name 17 Poecilia, M. picta and M. parae show similar expression of sex-linked genes in both sexes, 18 suggesting that dosage compensation has evolved. In contrast, although the Y chromosome of 19 their close relative, the guppy (Poecilia reticulata) is homologous to the Micropoecilia species' 20 X, it has not degenerated, but carries the same set of genes as the X chromosomes of all these 21 species (Figure 1). These findings suggest that Y-X recombination probably became suppressed 22 in a common ancestor of Micropoecilia, but in the guppy the Y occasionally recombines with 23 the X (Winge 1923), preventing degeneration. ...
... Under biologically plausible mutation rates, a Y-linked region can become highly 14 degenerated in several million generations in a population with an effective population size of 15 10,000, depending on the number of sites in the non-recombining genome region at which 16 deleterious mutations can occur; with 2,000 genes, if a single slightly deleterious mutation 17 renders a gene non-functional, it takes ~ 30 million generations for >90% of genes to be lost 18 (Bachtrog 2008). Degeneration is slower with fewer selected sites, and further gene loss after 19 10% of the ancestral genes have degenerated depends on spread of advantageous mutations, 20 and is extremely slow unless mutations are often advantageous. In larger populations, the 21 processes involved earlier in degeneration, Muller's ratchet and a reduced effective population 22 size due to selection eliminating deleterious mutations (see (Charlesworth 1996) are slower, 23 but the final stages are faster. ...
It is well known that the Y chromosomes of Drosophila and mammals, and the W chromosomes of birds carry only small fractions of the genes carried by the homologous X or Z chromosomes, and this "genetic degeneration"is associated with loss of recombination between the sex chromosome pair. However, it is still not known how much evolutionary time is needed to reach such nearly complete degeneration. The XY pair of species in a group of closely related poecilid fish are homologous, but have been found to have either non-degenerated, or completely degenerated, Y chromosomes. We evaluate evidence described in a recent paper, and show that the available data cast doubt on the view that degeneration has been extraordinarily rapid in the latter (Micropoecilia species).
... The simplest way to distinguish sex-linked loci from autosomal ones is to identify those found in reads that mapped to the sex chromosomes of the reference genome. However, this is not possible when (i) a reference genome is not available-as is the case for most wildlife species-and de novo genotyping is required, (ii) there is little conserved synteny between the studied genome and the reference, or (iii) the W/Y chromosome of the reference genome is fragmented into numerous unmapped scaffolds, as is common in many genome projects (Carvalho & Clark, 2013). ...
Identifying sex-linked markers in genomic datasets is important, because their analyses can reveal sex-specific biology, and their presence in supposedly neutral autosomal datasets can result in incorrect estimates of genetic diversity, population structure and parentage. But detecting sex-linked loci can be challenging, and available scripts neglect some categories of sex-linked variation. Here, we present new R functions to (1) identify and separate sex-linked loci in ZW and XY sex determination systems and (2) infer the genetic sex of individuals based on these loci. Two additional functions are presented, to (3) remove loci with artefactually high heterozygosity, and (4) produce input files for parentage analysis. We test these functions on genomic data for two sexually-monomorphic bird species, including one with a neo-sex chromosome system, by comparing biological inferences made before and after removing sex-linked loci using our function. We found that standard filters, such as low read depth and call rate, failed to remove up to 28.7% of sex-linked loci. This led to (i) overestimation of population FIS by ≤ 9%, and the number of private alleles by ≤ 8%; (ii) wrongly inferring significant sex-differences in heterozygosity, (iii) obscuring genetic population structure, and (iv) inferring ~11% fewer correct parentages. We discuss how failure to remove sex-linked markers can lead to incorrect biological inferences (e.g., sex-biased dispersal and cryptic population structure) and misleading management recommendations. For reduced-representation datasets with at least 15 known-sex individuals of each sex, our functions offer convenient, easy-to-use resources to avoid this, and to sex the remaining individuals.
... Taking a bespoke approach is critical as it reduces the phylogenetic distance between the sequence reads and the reference to which they are mapped, which can increase the proportion of reads that are accurately mapped and reduce issues arising from structural variation and repetitive sequence. Secondly, an important step in identifying diverged regions in sex chromosomes is ensuring stringent mapping parameters (Carvalho & Clark, 2013;Palmer et al., 2019;Smeds et al., 2015;. This is particularly relevant for homomorphic sex chromosomes as they still retain extensive sequence similarity between the X and Y, and incorrectly mapped reads can mask coverage differences between the sexes and lead to the misclassification of sex-linked sequences as autosomal. ...
Studies of sex chromosome systems at early stages of divergence are key to understanding the initial process and underlying causes of recombination suppression. However, identifying signatures of divergence in homomorphic sex chromosomes can be challenging due to high levels of sequence similarity between the X and the Y. Variations in methodological precision and underlying data can make all the difference between detecting subtle divergence patterns or missing them entirely. Recent efforts to test for X‐Y sequence differentiation in the guppy have led to contradictory results. Here, we apply different analytical methodologies to the same data set to test for the accuracy of different approaches in identifying patterns of sex chromosome divergence in the guppy. Our comparative analysis reveals that the most substantial source of variation in the results of the different analyses lies in the reference genome used. Analyses using custom‐made genome assemblies for the focal population or species successfully recover a signal of divergence across different methodological approaches. By contrast, using the distantly related Xiphophorus reference genome results in variable patterns, due to both sequence evolution and structural variations on the sex chromosomes between the guppy and Xiphophorus. Changes in mapping and filtering parameters can additionally introduce noise and obscure the signal. Our results illustrate how analytical differences can alter perceived results and we highlight best practices for the study of nascent sex chromosomes. Guppies, Poecilia reticulata, are at the extreme end of sex chromosome homomorphy. We use the same data across different methods and genome assemblies to determine the most sensitive approaches for detecting sex chromosomes.
... Напомним, что у чешуекрылых самцы гомогаметны (ZZ), а самки гетерогаметны (ZW или Z0). На основе выявления полоспецифической хромосомы Y основывались и методы определения пола малого булавоусого хрущака Tribolium castaneum (Lagisz et al., 2010б), комаров рода Anopheles (Krzywinski et al., 2004;Hall et al., 2013), клопа Rhodnius proximus (Koerich et al., 2016), плодовой мушки Drosophila melanogaster (Carvalho and Clark, 2013). При этом подходе подразумевается, что в отсутствии полоспецифичной хромосомы (Y или W) отсутствует и ПЦР-продукт. ...
Insects play an important role in biocenoses due to their abundance and wide (cosmopolitan) distribution. Many insects are crop pests. An effective pest control could be realized in case of proper species identification, which is usually managed by morphological analysis. Molecular methods allow to deep study of many issues of insect biology. In particular, traditional approach can not ordinary identify a species at all stages of their life cycle, whereas molecular methods can it. This review covers a wide range of issues related to the molecular genetic analysis of insects. In the first section we consider the methods of fixation and storage of insect specimens, as well as their impact on DNA quality. Further, we provide general information on population study design. Various schemes of DNA extraction, examples of both express techniques and more thorough protocols for DNA extraction and their purification are provided. In addition, methods of DNA isolation that allow to preserve a specimen integrity for further morphological studies are considered. The methods of DNA quality control are described in detail, that is important for PCR analysis. The last section provides various methods of PCR analysis, that we exemplify by studies aimed to elucidate both fundamental issues and practical problems.
... Il est cependant possible de récupérer les lectures d'individus hétérogamétiques n'ayant pas été alignées puis de les assembler pour reconstruire les séquences Y-spécifiques. Enfin, il est possible d'utiliser une approche dite des k-mers (voir Carvalho et Clark, 2013). Elle consiste à fragmenter les génomes mâles et femelles en fragments de longueurs k puis d'identifier les fragments dont les ratios de couverture f emelles/mâles diffèrent de 1. Les k-mers spécifiques du chromosome Y auront un ratio de couverture f emelles/mâles de 0, les k-mers des séquences X auront un ratio de couverture f emelles/mâles de 2. ...
La trajectoire décrivant l’évolution d’une paire de chromosomes sexuels à longtemps été proposée comme étant universelle pour tous les systèmes, cependant des propositions alternatives ont récemment nuancé ce «modèle» unique. D’après ce modèle, il y aurait dans un premier temps l’émergence d’une région non-recombinante (XY ou ZW), puis, une expansion de celle-ci. Simultanément, l’absence de recombinaison induit ce que l’on appelle la dégénérescence du chromosome Y (ou W). La dégénérescence est supposée augmenter et, après un certain temps évolutif, devrait conduire à un système dans lequel le chromosome Y (ou W) serait plus petit que le chromosome X (ou Z), voire disparaît. Cependant, seulement une trentaine de paires de chromosomes sexuels de plantes ont été étudiées avec des données empiriques, parmi plus de 15 000 espèces dioïques (i.e. plantes à sexes séparés). Il en résulte que certaines étapes du systèmes sont mieux supportées que d’autres. Plus précisément, la formation de la région non-recombinante a essentiellement été étudiée de manière théorique, tandis qu’une forte dégénérescence avec un chromosome Y (ou W) plus petit que le chromosome X (ou Z) n’a été décrite que chez les animaux. Afin de mieux décrire la première étape du modèle, l’émergence de la région non recombinante, le premier axe de cette thèse représente une étude de Silene acaulis ssp exscapa, la seule sous-espèce dioïque du complexe Silene acaulis. En effet, ceci laisse supposer que ce système sexuel est un caractère dérivé, donc probablement récent. Le mécanisme du déterminisme du sexe n’étant pas connu, j’ai voulu savoir si une région non-recombinante typique d’une paire de chromosomes sexuels est présente chez cette sous-espèce. Pour cela, j’ai utilisé un outil récemment publié basé sur l’analyse de fréquences génotypiques et phénotypiques de mâles et de femelles au sein d’une population. Deux jeux de données RNA-seq provenant de deux populations différentes ont permis d’identifier 27 gènes potentiellement XY, et suggèrent que la paire de chromosomes sexuels serait récente. Des analyses complémentaires sont tout de même nécessaires pour confirmer ces résultats. Deuxièmement, afin de tester l’existence d’une paire ancienne de chromosomes sexuels avec une forte dégénérescence chez les plantes, le deuxième axe de cette thèse est une étude de deux espèces dioïques de la famille des Cannabaceae, Cannabis sativa et Humulus lupulus. En effet, l’ancêtre commun de ces deux espèces, qui divergent depuis plusieurs dizaines de millions d’années, était probablement dioïque. De plus, des analyses cytologiques ont identifié des paires de chromosomes sexuels qui pourraient être anciennes. Pour caractériser l’âge et le niveau de dégénérescence de ces paires de chromosomes sexuels, des données RNA-seq d’un croisement ont été générées pour chacune des deux espèces. Un outil probabiliste analysant les ségrégations alléliques au sein d’un croisement a permis d’identifier la première paire de chromosomes sexuels homologue entre deux genres chez les plantes. De plus, ces chromosomes sexuels sont parmi les plus vieux et les plus dégénérés actuellement décrits chez les plantes. Par ailleurs, la détection de séquence Y-spécifiques pourrait permettre d’améliorer la culture de ces deux espèces puisque seules les femelles ont un intérêt économique et que le dimorphisme sexuel est faible. J’ai développé des amorces PCR qui montrent des résultats prometteurs. Plus généralement, ces résultats apportent de nouvelles informations concernant les étapes les moins bien décrites de l’évolution des chromosomes sexuels chez les plantes. Premièrement, nous montrons qu’une paire de chromosomes sexuels a probablement émergé récemment dans une espèce, et confirmons l’intérêt de continuer à l’étudier. Deuxièmement, nous confirmons que des chromosomes sexuels vieux et fortement dégénérés existent chez les plantes.
... Finally, older strata of Y or W chromosome could be detected using a k-mer approach, by matching short reads of the homogametic sex to the genome of the heterogametic sex (Carvalho & Clark, 2013), or using in silico whole genome subtraction (Dissanayake et al., 2020). ...
Sex‐specific ecology has management implications, but rapid sex‐chromosome turnover in fishes hinders sex‐marker development for monomorphic species. We used annotated genomes and reduced‐representation sequencing data for two Australian percichthyids, Macquarie perch Macquaria australasica and golden perch M. ambigua, and whole genome resequencing for 50 Macquarie perch of each sex, to identify sex‐linked loci and develop an affordable sexing assay. In‐silico pool‐seq tests of 1,492,004 Macquarie perch SNPs revealed that a 275‐Kb scaffold was enriched for gametologous loci. Within this scaffold, 22 loci were sex‐linked in a predominantly XY system, with females being homozygous for the X‐linked allele at all 22, and males having the Y‐linked allele at >7. Seven XY‐gametologous loci (all males, but no females, are heterozygous or homozygous for the male‐specific allele) were within a 146‐bp region. A PCR‐RFLP sexing assay targeting one Y‐linked SNP, tested in 66 known‐sex Macquarie perch and two of each sex of three confamilial species, plus amplicon sequencing of 400 bp encompassing the 146‐bp region, revealed that the few sex‐linked positions differ between species and between Macquarie perch populations. This indicates sex‐chromosome lability in Percichthyidae, supported by non‐homologous scaffolds containing sex‐linked loci for Macquarie‐ and golden perches. The present resources facilitate genomic research in Percichthyidae, including formulation of hypotheses about candidate genes of interest such as transcription factor SOX1b that occurs in the 275‐Kb scaffold ~38 Kb downstream of the 146‐bp region containing seven XY‐gametologous loci. Sex‐linked markers will be useful for determining genetic sex in some populations and studying sex chromosome turnover.
... We used the YGS method to computationally identify the Y-linked scaffolds by comparing the assembled genome with male and female Illumina reads [13]. This procedure identifies Y-linked sequences but not the genes they contain. ...
... This procedure identifies Y-linked sequences but not the genes they contain. For example, in D. virilis, most Y-linked scaffolds contain only repetitive DNA and other non-coding sequences [13]. In order to identify and annotate the protein-encoding genes we did blast searches in the Y-linked sequences identified by the YGS method against databases of Dipteran proteins, bacterial and yeast proteins (to identify contaminant scaffolds), and transposable elements. ...
... The bimodal distribution separates the Y-linked sequences (peak on the right-hand, with a large amount of sequence unmatched by female reads) from the X and autosomes (left-hand peak). The red dots mark the scaffolds matching the eight previously known Y-linked genes of D. willistoni [13,14]. All these sequences are located in the right-hand peak (average for Sanger assembly: 94%; range: 87-100%); they served as a positive control and show that the YGS method reliably identifies Y-linked sequences in D. willistoni. ...
Y chromosomes play important roles in sex determination and male fertility. In several groups (e.g., mammals) there is strong evidence that they evolved through gene loss from a common X-Y ancestor, but in Drosophila the acquisition of new genes plays a major role. This conclusion came mostly from studies in two species. Here we report the identification of the 22 Y-linked genes in D. willistoni. They all fit the previously observed pattern of autosomal or X-linked testis-specific genes that duplicated to the Y. The ratio of gene gains to gene losses is ~25 in D. willistoni, confirming the prominent role of gene gains in the evolution of Drosophila Y chromosomes. We also found four large segmental duplications (ranging from 62 kb to 303 kb) from autosomal regions to the Y, containing ~58 genes. All but four of these duplicated genes became pseudogenes in the Y or disappeared. In the GK20609 gene the Y-linked copy remained functional, whereas its original autosomal copy degenerated, demonstrating how autosomal genes are transferred to the Y chromosome. Since the segmental duplication that carried GK20609 contained six other testis-specific genes, it seems that chance plays a significant role in the acquisition of new genes by the Drosophila Y chromosome.
... In contrast, in older strata with substantial Y or W degeneration, X-and Z-linked loci will be effectively hemizygous in the heterogametic sex (halved read depth), and higher SNP density is expected in the homogametic sex. Finally, older strata of Y or W chromosome could be detected using a k-mer approach, by matching short reads of the homogametic sex to the genome of the heterogametic sex (Carvalho & Clark, 2013), or usingin-silico whole genome subtraction (Dissanayake et al., 2020). A combination of different approaches might be needed to detect sex-linked loci when the age of sex chromosomes is unknown. ...
Sex-specific ecology has management implications, but rapid sex-chromosome turnover in fishes hinders development of markers to sex monomorphic species. Here, we use annotated genomes and reduced-representation sequencing data for two Australian percichthyids, the Macquarie perch Macquaria australasica and the golden perch M. ambigua, and whole genome resequencing data for 50 Macquarie perch of each sex, to detect sex-linked loci, identify a candidate sex-determining gene and develop an affordable sexing assay. In-silico pool-seq tests of 1,492,004 Macquarie perch SNP loci revealed that a 275-Kb scaffold, containing the transcription factor SOX1b gene, was enriched for gametologous loci. Within this scaffold, 22 loci were sex-linked in a predominantly XY system, with females being homozygous at all 22, and males being heterozygous at two or more. Seven XY-gametologous loci were within a 146-bp region. Being ~38 Kb upstream of SOX1b, it might act as an enhancer controlling SOX1b transcription in the bipotential gonad that drives gonad differentiation. A PCR-RFLP sexing assay, targeting one of the Y-linked SNPs, tested in 66 known-sex Macquarie perch and two individuals of each sex of three confamilial species, and amplicon sequencing of 400 bp encompassing the 146-bp region, revealed that the few sex-linked positions differ between species and between Macquarie perch populations. This indicates sex-chromosome lability in Percichthyidae, also supported by non-homologous scaffolds containing sex-linked loci for Macquarie- and golden perches. The resources developed here will facilitate genomic research in Percichthyidae. Sex-linked markers will be useful for determining genetic sex in some populations and studying sex chromosome turnover.
