Assessment of 17 microsatellite loci for their use in parentage verification and individual identification in the Balkan donkey breed

Stanišić, Ljubodrag; Dimitrijević, Vojin; Simeunović, Predrag; Glavinić, Uroš; Jovanović, Biljana; Stevanović, Jevrosima; Stanimirović, Zoran

doi:10.2298/gensr1701021s

Cited by 3 publications

(9 citation statements)

References 11 publications

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“…The COR022 and LEX73 loci failed to amplify in all the samples and did not provide data for reliable analysis while the ASB17 marker was monomorphic (91 bp) for all examined animals. Similar results were reported in the Balkan donkey populations [ 35 ]. This study is the first systematic large-scale study of different geographical regions populations from Turkey by using the microsatellites.…”

Section: Discussionsupporting

confidence: 91%

Genetic Characterization of Native Donkey (Equus asinus) Populations of Turkey Using Microsatellite Markers

Yatkın

Özdil

Ünal

et al. 2020

Animals

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This study presents the first insights to the genetic diversity and structure of the Turkish donkey populations. The primary objectives were to detect the main structural features of Turkish donkeys by microsatellite markers. A panel of 17 microsatellite markers was applied for genotyping 314 donkeys from 16 locations of Turkey. One hundred and forty-two alleles were identified and the number of alleles per locus ranged from 4 to 12. The highest number of alleles was observed in AHT05 (12) and the lowest in ASB02 and HTG06 (4), while ASB17 was monomorphic. The mean HO in the Turkish donkey was estimated to be 0.677, while mean HE was 0.675. The polymorphic information content (PIC) was calculated for each locus and ranged from 0.36 (locus ASB02) to 0.98 (locus AHT05), which has the highest number of alleles per locus in the present study. The average PIC in our populations was 0.696. The average coefficient of gene differentiation (GST) over the 17 loci was 0.020 ± 0.037 (p < 0.01). The GST values for single loci ranged from −0.004 for LEX54 to 0.162 for COR082. Nei’s gene diversity index (Ht) for loci ranged from 0.445 (ASB02) to 0.890 (AHT05), with an average of 0.696. A Bayesian clustering method, the Structure software, was used for clustering algorithms of multi-locus genotypes to identify the population structure and the pattern of admixture within the populations. When the number of ancestral populations varied from K = 1 to 20, the largest change in the log of the likelihood function (ΔK) was when K = 2. The results for K = 2 indicate a clear separation between Clade I (KIR, CAT, KAR, MAR, SAN) and Clade II (MAL, MER, TOK, KAS, KUT, KON, ISP, ANT, MUG, AYD and KAH) populations.

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Section: Discussionsupporting

confidence: 91%

Genetic Characterization of Native Donkey (Equus asinus) Populations of Turkey Using Microsatellite Markers

Yatkın

Özdil

Ünal

et al. 2020

Animals

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“…In conclusion, compared to the horse‐specific multiplex systems, which have been previously used to investigate parentage and genetic diversity in donkeys , our new donkey‐specific 13‐plex typing system has six advantages: 1) our system contains five additional STR loci recommended by the ISAG, which were previously unavailable; 2) redesigned primers solve issues with nonspecific amplification and unsuitability of the multiplex system; 3) our system includes three fluorescent dyes, thus primers for additional STR loci can potentially be added to this system by designing allele products with fragment sizes that differ from those of other existing STR primers, or by labelling them with different fluorescent dyes; 4) uses the low‐cost polymerase 2xEasyTaq PCR Supermix, which can efficiently obtain excellent genotype profiles; 5) PCR reaction can be completed within 73 min, which is faster than those in previous studies (84–144 min) ; (6) system provides stutter data for the first time in donkeys. These advantages should help researchers obtain more confident genotyping results with STRs in donkeys.…”

Section: Discussionmentioning

confidence: 99%

“…The genetic diversity of Catalonian and Spanish donkeys has been investigated for several decades , where the StockMarks for Horses had been used. In 2017, the Equine Genotypes Panel 1.1, which contains primers for 17 horse STR loci, was used to verify the parentage and identify individuals within the endangered Balkan donkey . However, primers for three of the horse STR loci (ASB2, ASB17 and HMS1) were unsuitable for the donkey, and thus, had to be excluded from the analysis .…”

Section: Introductionmentioning

confidence: 99%

“…In 2017, the Equine Genotypes Panel 1.1, which contains primers for 17 horse STR loci, was used to verify the parentage and identify individuals within the endangered Balkan donkey . However, primers for three of the horse STR loci (ASB2, ASB17 and HMS1) were unsuitable for the donkey, and thus, had to be excluded from the analysis . In this same year, the International Society for Animal Genetics (ISAG) recommended the use of 13 autosomal dinucleotide STRs (for the loci AHT4, ASB23, HMS2, HMS3, HMS6, HMS7, HMS18, HTG7, HTG10, TKY297, TKY312, TKY337 and TKY343) for individual identification and parentage testing in the donkey .…”

Section: Introductionmentioning

confidence: 99%

“…This was the first time that the four TKY loci and HMS18 had been recommended for donkey parentage verification. Of the 13 recommended loci, five (HMS18, TKY297, TKY312, TKY337 and TKY343) are not included in the horse panel , and, furthermore, due to differences in the genomic sequences between donkey and horse at the primer sites, amplification of alleles at several other STR loci is inefficient or fails, thus limiting their usefulness . Currently, in the literature, there is no reported donkey‐specific multiplex typing system for all 13 recommended STRs; producing one should help improve the accuracy and efficiency of individual identification, paternity testing and genetic analysis of populations.…”

Section: Introductionmentioning

confidence: 99%

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A novel 13‐plex STR typing system for individual identification and parentage testing of donkeys (Equus asinus)

Dang

Shang

Zhang

et al. 2019

Equine Veterinary Journal

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Summary Background Previous studies investigating donkey parentage and genetic diversity used horse‐specific multiplex systems. However, several mis‐allele and null‐allele issues were found with some of the horse primers when used in donkeys. In 2017, the International Society for Animal Genetics (ISAG) recommended 13 dinucleotide short tandem repeats (STRs) (AHT4, ASB23, HMS2, HMS3, HMS6, HMS7, HMS18, HTG7, HTG10, TKY297, TKY312, TKY337 and TKY343) as a core panel that should be used to identify individuals and to test for parentage in donkeys. To date, no single multiplex STR typing system containing all 13 donkey STRs recommended by the ISAG has been reported. Objectives To establish a novel and donkey‐specific multiplex STR typing system containing all 13 recommended STRs. Study design Assay development and validation in field population. Methods Primers for seven of the STRs were redesigned and conditions for polymerase chain reaction (PCR) were optimised. We analysed the allele sequences, sensitivity, species‐specificity and stutter ratios of this new system. Results A 13‐plex STR typing system for donkey was established. A full profile could be generated from a single PCR reaction using as little as 5 ng of DNA template with the 13 pairs of primers labelled with fluorescent dyes. An allele ladder, containing 101 alleles from the 13 STRs, was generated. No full genotype profile was generated with these primers if DNA from humans, or 11 other commonly encountered animals, was used. Genotypes could be generated for the horse and horse‐donkey hybrids (mule and hinny). Stutter ratios and population genetic parameters were calculated based on samples from 150 donkeys. The combined probabilities of paternity exclusion for this system were 0.988907326 (CPEduo) and 0.999665018 (CPEtrio). Main limitations This system cannot detect sex. Conclusions Our results indicate that our donkey‐specific 13‐plex STR typing system is sensitive, species‐specific and robust for individual identification, paternity testing and population genetic analysis in donkeys, and has potential forensic applications.

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Morphological and genetic characterization of the Graciosa donkey breed

Lopes

Azevedo

Mendonça

et al. 2023

Journal of Applied Animal Research

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Assessment of 17 microsatellite loci for their use in parentage verification and individual identification in the Balkan donkey breed

Abstract: The aim of this study was to assess a panel of 17 microsatellites for parentage verification and individual identification in the endangered Balkan donkey breed. Allele frequencies for 17 microsatellite loci (AHT4,

Cited by 3 publications

References 11 publications

Genetic Characterization of Native Donkey (Equus asinus) Populations of Turkey Using Microsatellite Markers

Genetic Characterization of Native Donkey (Equus asinus) Populations of Turkey Using Microsatellite Markers

A novel 13‐plex STR typing system for individual identification and parentage testing of donkeys (Equus asinus)

Morphological and genetic characterization of the Graciosa donkey breed

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