Using blood and non-invasive shed skin samples to identify sex of caenophidian snakes based on multiplex PCR assay

Tawichasri, Panupong; Laopichienpong, Nararat; Chanhome, Lawan; Phatcharakullawarawat, Rattanin; Singchat, Worapong; Koomgun, Tassica; Prasongmaneerut, Tulyawat; Rerkamnuaychoke, Wuttiporn; Sillapaprayoon, Siwapech; Muangmai, Narongrit; Suntrarachun, Sunutcha; Baicharoen, Sudarath; Peyachoknagul, Surin; Srikulnath, Kornsorn

doi:10.1016/j.jcz.2017.11.003

Cited by 13 publications

(18 citation statements)

References 34 publications

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“…All cobras were exposed to ambient light cycles. The sex of each individual was identified morphologically by mating observations and sexing probes that searched for the male hemipenes (Laszlo, 1975), and confirmed by a molecular sexing approach using specific codominant and dominant DNA markers based on gametologous genes (Laopichienpong, Muangmai, et al, 2017;Laopichienpong, Tawichasri, et al, 2017;Tawichasri et al, 2017). Animal care and all experimental procedures were approved by the Animal Experiment Committee, Kasetsart University (approval no.…”

Section: Specimen Collection and Dna Extractionmentioning

confidence: 99%

“…Snakes mostly exhibit genotypic sex determination for ZZ/ZW type (Laopichienpong, Muangmai, et al, 2017;Laopichienpong, Tawichasri, et al, 2017;Tawichasri et al, 2017), and telomere dynamics have been characterized in snake species including water pythons (Liasis fuscus), garter snakes (Thamnophis elegans), and red-sided garter snakes (Thamnophis sirtalis parietalis) (Bronikowski, 2008;Rollings et al, 2017;Ujvari & Madsen, 2009). The present study makes use of precise individual age records from actively maintained populations of Siamese cobra (Naja kaouthia) to improve our understanding of age-and sex-related telomere dynamics in ZZ/ ZW species.…”

Section: Introductionmentioning

confidence: 99%

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Dynamics of telomere length in captive Siamese cobra (Naja kaouthia) related to age and sex

Singchat

Kraichak

Tawichasri

et al. 2019

Ecology and Evolution

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Telomeres comprise tandem repeated DNA sequences that protect the ends of chromosomes from deterioration or fusion with neighboring chromosomes, and their lengths might vary with sex and age. Here, age‐ and sex‐related telomere lengths in male and female captive Siamese cobras (Naja kaouthia) were investigated using quantitative real‐time polymerase chain reaction based on cross‐sectional data. A negative correlation was shown between telomere length and body size in males but not in females. Age‐related sex differences were also recorded. Juvenile female snakes have shorter telomeres relative to males at up to 5 years of age, while body size also rapidly increases during this period. This suggests that an accelerated increase in telomere length of female cobra results from sex hormone stimulation to telomerase activity, reflecting sexually dimorphic phenotypic traits. This might also result from amplification of telomeric repeats on sex chromosomes. By contrast, female Siamese cobras older than 5 years had longer telomeres than males. Diverse sex hormone levels and oxidative stress parameters between sexes may affect telomere length.

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Section: Specimen Collection and Dna Extractionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Dynamics of telomere length in captive Siamese cobra (Naja kaouthia) related to age and sex

Singchat

Kraichak

Tawichasri

et al. 2019

Ecology and Evolution

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“…It is also the only method capable of identifying the sex of embryos and juveniles before the development of primary and secondary sexual characteristics (Morinha et al, 2012). Sex chromosomes are conserved in birds, and the majority of mammals and snakes; therefore, established molecular sexing techniques are often applied throughout these groups (Fridolfsson and Ellegren, 1999;Fernando and Melnick, 2001;Morin et al, 2005;Brubaker et al, 2011;Morinha et al, 2012Morinha et al, , 2015Laopichienpong et al, 2017a,b;Tawichasri et al, 2017). However, no such widely applicable technique was thought possible for other amniotes because they show variability in sexdetermination systems and/or homomorphic sex chromosomes (Ezaz et al, 2009b;Pokorná and Kratochvíl, 2009;Gamble, 2010;Sarre et al, 2011;Gamble et al, 2015).…”

Section: How Many Functional Gene Markers Are Required For Sex Identimentioning

confidence: 99%

“…The use of a small number of genes to identify sex determination and sex-chromosomal linkages might result in errors because many homologous loci across amniote sex chromosomes were observed at Y-specific loci in the current study, while false positive qPCR results might often be obtained in different laboratories (Rovatsos et al, 2014b). Developmental molecular identification and analysis of sex-linked genetic markers based on a PCR approach are required to establish a genotyping tool for practical sexing of individuals in a commercial breeding program and allow accurate determination of sex in populations through insight into the evolutionary history of the Y chromosome (or the W chromosome in a female heterogametic species, such as occurs in snakes) (Laopichienpong et al, 2017a,b;Tawichasri et al, 2017).…”

Section: How Many Functional Gene Markers Are Required For Sex Identimentioning

confidence: 99%

Genome Complexity Reduction High-Throughput Genome Sequencing of Green Iguana (Iguana iguana) Reveal a Paradigm Shift in Understanding Sex-Chromosomal Linkages on Homomorphic X and Y Sex Chromosomes

et al. 2020

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The majority of lizards classified in the superfamily Iguanoidea have an XX/XY sexdetermination system in which sex-chromosomal linkage shows homology with chicken (Gallus gallus) chromosome 15 (GGA15). However, the genomics of sex chromosomes remain largely unexplored owing to the presence of homomorphic sex chromosomes in majority of the species. Recent advances in high-throughput genome complexity reduction sequencing provide an effective approach to the identification of sexspecific loci with both single-nucleotide polymorphisms (SNPs) and restriction fragment presence/absence (PA), and a better understanding of sex chromosome dynamics in Iguanoidea. In this study, we applied Diversity Arrays Technology (DArTseq TM) in 29 phenotypic sex assignments (14 males and 15 females) of green iguana (Iguana iguana). We confirmed a male heterogametic (XX/XY) sex determination mode in this species, identifying 29 perfectly sex-linked SNP/PA loci and 164 moderately sex-linked SNP/PA loci, providing evidence probably indicative of XY recombination. Three loci from among the perfectly sex-linked SNP/PA loci showed partial homology with several amniote sex chromosomal linkages. The results support the hypothesis of an ancestral super-sex chromosome with overlaps of partial sex-chromosomal linkages. However, only one locus among the moderately sex-linked loci showed homology with GGA15, which suggests that the specific region homologous to GGA15 was located outside the non-recombination region but in close proximity to this region of the sex chromosome

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“…Such risk-free approaches lend themselves especially to citizen science projects, where members of the public can collect and ship samples from a far larger area and at far lower cost. DNA derived from shed skins of lizards and snakes has long been known to be of sufficient quality and quantity for PCR-based genotyping of a small number of genetic markers (Bricker, Bushar, Reinert, & Gelbert, 1996; Fetzner Jr, 1999; Horreo, Peláez, & Fitze, 2015; Tawichasri et al, 2017; Villarreal, Bricker, Reinert, Gelbert, & Bushar, 1996), and so here we assess the utility of shed skin for larger-scale single nucleotide polymorphism (SNP) genotyping using genotyping-by-sequencing (GBS).…”

Section: Introductionmentioning

confidence: 99%

Shed skin as a source of DNA for genotyping-by-sequencing (GBS) in reptiles

Brekke

Shier

Hegarty

et al. 2019

Preprint

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Association and genetic mapping studies aimed at linking genotype to phenotype are powerful tools that require large numbers of samples, complicating their use in long-lived species with low fecundity. Shed skins of snakes and other reptiles contain DNA; are a safe and ethical way of non-invasively sampling large numbers of individuals; and provide a simple mechanism by which to involve the public in scientific research. Here we test whether the DNA in dried shed skins mailed to us from citizen scientists is suitable for reduced representation sequencing approaches, specifically genotyping-by-sequencing (GBS). We find that shed skin samples provide DNA of sufficient quality and quantity for GBS, although libraries from shed skin resulted in fewer sequenced reads than libraries from snap-frozen muscle, and contained slightly fewer variants (70,685 SNPs versus 97,724). This issue is a direct result of lower read counts of the shed skin samples, and can be rectified quite simply with deeper sequencing. Skin-derived libraries also have a very slight (but significantly different) profile of transitions and transversions, suggesting that DNA damage occurs but is minimal. We conclude that shed skin-derived DNA is a good source of genomic DNA for a variety of genetic studies, and use it to identify sex-linked scaffolds in the corn snake genome.

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Using blood and non-invasive shed skin samples to identify sex of caenophidian snakes based on multiplex PCR assay

Cited by 13 publications

References 34 publications

Dynamics of telomere length in captive Siamese cobra (Naja kaouthia) related to age and sex

Dynamics of telomere length in captive Siamese cobra (Naja kaouthia) related to age and sex

Genome Complexity Reduction High-Throughput Genome Sequencing of Green Iguana (Iguana iguana) Reveal a Paradigm Shift in Understanding Sex-Chromosomal Linkages on Homomorphic X and Y Sex Chromosomes

Shed skin as a source of DNA for genotyping-by-sequencing (GBS) in reptiles

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