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Home / Equine Research Bank / Anim Genet / Identification of processed pseudogenes in the genome of Tho...
Animal genetics2022; 53(2); 183-192; doi: 10.1111/age.13174

Identification of processed pseudogenes in the genome of Thoroughbred horses: Possibility of gene-doping detection considering the presence of pseudogenes.

Abstract: Processed pseudogenes, also known as retrocopy genes, are copies of messenger RNAs that have been reverse transcribed into DNA and inserted into the genome. In this study, we identified 62 processed pseudogene candidates as intron-less genes from whole-genome sequencing (WGS) data of Thoroughbred horses using delly structural variation software. The 62 processed pseudogene candidates were confirmed by PCR amplification of intron-less products. A total of 11 processed pseudogenes were confirmed in the genome of all 23 analysed horses, whereas three processed pseudogenes with structures of ATP11B, DPH3 and RPL17 were detected in only one of 115 horses by PCR amplification of intron-less products. Currently, most of the gene doping tests proposed in human and horse sports are adapted PCR-based methods using hydrolysis probes to detect exon/exon junctions in transgenes because the operation is simple and economical. However, when the pseudogene is present in the host genome, the PCR-based methods may have a potential risk of detecting false positives. In this study, because processed pseudogenes that exist less frequently in the horse genome may affect PCR-based transgene detection in gene-doping tests, we propose and demonstrate that PCR amplification and sequencing using primers designed on transgene and promotors and/or polyadenylation signal for gene expression are useful for gene-doping detection as an additional confirmatory test to prevent false positives.
© 2022 Stichting International Foundation for Animal Genetics.
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Publication Date: 2022-01-25 PubMed ID: 35077588DOI: 10.1111/age.13174Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research conducted a thorough analysis of the Thoroughbred horses’ genome, revealing the presence of 62 processed pseudogenes (intron-less genes). The study, beyond its genetic insights, tackles the potential issue of false positives in gene-doping detection, suggesting a more accurate PCR-based method considering pseudogenes.

Overview of the Research

  • The research examined the presence of processed pseudogenes within the genome of Thoroughbred horses. These pseudogenes are variants of regular genes, which have been reverse transcribed into DNA and inserted back into the genome. Conceptually, they can be thought of as ‘false’ genes or ‘non-functioning’ copies of existing genes in the organism.
  • The researchers identified these pseudogenes via whole-genome sequencing (WGS) data. Using delly structural variation software, they found 62 potential processed pseudogenes. These candidates were then confirmed through PCR (Polymerase Chain Reaction) amplification of intron-less products – a process that stimulates the replication of specific DNA segments.

Significance of Pseudogenes in Gene-Doping Detection

  • The team explored how these pseudogenes might influence gene-doping tests in sports, both human and horse-based. Many of the tests that currently detect gene-doping utilise PCR-based methods. This type of test uses hydrolysis probes to detect exon/exon junctions in transgenes. It’s a popular method due to its simplicity and cost-effectiveness.
  • However, the presence of pseudogenes can potentially result in false positives during the doping tests. If a test picks up on the presence of these ‘non-functioning’ genes, it might incorrectly identify them as the product of gene-doping.

A Proposed Solution

  • The researchers suggest an alternative protocol for doping tests to avoid these false readings. The gear of focus is on utilising specific primers during PCR amplification and sequencing. These primers should be designed on the transgene, promoters, and/or polyadenylation signals for gene expression.
  • Through this method, the test can focus more clearly on the substances indicative of gene-doping, while ignoring the pseudogenes that could otherwise lead to false positives. Essentially, these new guidelines add an extra layer of scrutiny to prevent the risk of incorrect results.

Cite This Article

APA
Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Kusano K, Nagata SI. (2022). Identification of processed pseudogenes in the genome of Thoroughbred horses: Possibility of gene-doping detection considering the presence of pseudogenes. Anim Genet, 53(2), 183-192. https://doi.org/10.1111/age.13174

Publication

Animal genetics
ISSN: 1365-2052
NlmUniqueID: 8605704
Country: England
Language: English
Volume: 53
Issue: 2
Pages: 183-192

Researcher Affiliations

Tozaki, Teruaki
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Ohnuma, Aoi
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Kikuchi, Mio
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Ishige, Taichiro
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Kakoi, Hironaga
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Hirota, Kei-Ichi
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.
Kusano, Kanichi
  • Equine Department, Japan Racing Association, Minato, Tokyo, Japan.
Nagata, Shun-Ichi
  • Genetic Analysis Department, Laboratory of Racing Chemistry, Utsunomiya, Tochigi, Japan.

MeSH Terms

  • Animals
  • DNA Primers
  • Doping in Sports
  • Genome
  • Horses / genetics
  • Introns
  • Pseudogenes

Grant Funding

  • Japan Racing Association

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Citations

This article has been cited 4 times.
  1. Lu Y, Yan J, Ou G, Fu L. A Review of Recent Progress in Drug Doping and Gene Doping Control Analysis.. Molecules 2023 Jul 18;28(14).
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  2. Batcher K, Varney S, Raudsepp T, Jevit M, Dickinson P, Jagannathan V, Leeb T, Bannasch D. Ancient segmentally duplicated LCORL retrocopies in equids.. PLoS One 2023;18(6):e0286861.
    doi: 10.1371/journal.pone.0286861pubmed: 37289743google scholar: lookup
  3. Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Takahashi Y, Nagata SI. Short Insertion and Deletion Discoveries via Whole-Genome Sequencing of 101 Thoroughbred Racehorses.. Genes (Basel) 2023 Mar 3;14(3).
    doi: 10.3390/genes14030638pubmed: 36980910google scholar: lookup
  4. Tozaki T, Ohnuma A, Nakamura K, Hano K, Takasu M, Takahashi Y, Tamura N, Sato F, Shimizu K, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Hamilton NA, Nagata SI. Detection of Indiscriminate Genetic Manipulation in Thoroughbred Racehorses by Targeted Resequencing for Gene-Doping Control.. Genes (Basel) 2022 Sep 4;13(9).
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