PCR-RFLP assays to detect recessive lethal alleles in Landrace and Duroc pigs in Vietnam

Abstract

Increasing the number of live-born piglets per litter is one of the major objectives of pig breeding programs. Single nucleotide polymorphisms (SNPs) in polynucleotide kinase-phosphatase (PNKP), RNA polymerase I subunit B (POLR1B), ribosome biogenesis (URB1), and transcriptional adaptors ADA2 (TADA2A genes are recessive lethal alleles that are associated with embryonic lethality, which affects the reproductive performance and animal welfare of pig populations). No assay is available to screen for the recessive haplotypes of these 4 SNPs in pig populations. We successfully developed and verified a rapid, simple, and cost-effective PCR–restriction-fragment length polymorphism (PCR-RFLP) assay for genotyping SNPs in PNKP, POLR1B, URB1 genes in Landrace and TADA2A genes in Duroc pigs in Vietnam. Our PCR-RFLP approach could potentially be applied in screening programs on a larger scale to improve the reproductive performance of commercial pig populations.

Keywords

Duroc Landrace PCR-RFLP SNP swine

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References

Ahmadian

, et al. Pyrosequencing: history, biochemistry and future. Clin Chim Acta 2006;363:83–94.

Alavian

, et al. A facile PCR-RFLP method for genotyping of ITPA rs1127354 and rs7270101 polymorphisms. J Clin Lab Anal 2018;32:e22440.

Beuzen

, et al. Molecular markers and their use in animal breeding. Vet J 2000;160:42–52.

Bidarimath

Tayade

Pregnancy and spontaneous fetal loss: a pig perspective. Mol Reprod Dev 2017;84:856–869.

Boushaba

Tabet-Aoul

Allele and genotype frequencies of β-lactoglobulin gene using PCR-RFLP in Algerian local cattle populations. Mol Biol Res Commun 2024;13:43–49.

Derks

MFL

, et al. Balancing selection on a recessive lethal deletion with pleiotropic effects on two neighboring genes in the porcine genome. PLoS Genet 2018;14:e1007661.

Derks

MFL

, et al. Loss of function mutations in essential genes cause embryonic lethality in pigs. PLoS Genet 2019;15:e1008055.

Griffin

Smith

LM.

Single-nucleotide polymorphism analysis by MALDI-TOF mass spectrometry. Trends Biotechnol 2000;18:77–84.

Keramati

, et al. In silico design and in vitro validation of a novel PCR-RFLP assay for determination of phylogenetic clusters of streptokinase gene alleles in streptococci groups. Microb Pathog 2020;139:103862.

10.

Montanez-Gonzalez

, et al. A PCR-RFLP method for genotyping of inversion 2Rc in Anopheles coluzzii. Parasit Vectors 2021;14:174.

11.

Ordovas

JM.

Separation of small-size DNA fragments using agarose gel electrophoresis. Methods Mol Biol 1998;110:35–42.

12.

Paek

H-J

, et al. Application of PCR-RFLP for quick identification of MSTN mutants in MSTN mutant pig breeding. Anim Biotechnol 2023;34:2231–2239.

13.

Pankotai

, et al. The homologous Drosophila transcriptional adaptors ADA2a and ADA2b are both required for normal development but have different functions. Mol Cell Biol 2005;25:8215–8227.

14.

Putnová

, et al. A new HpaII PCR-RFLP within the porcine prolactin receptor (PRLR) gene and study of its effect on litter size and number of teats. J Anim Breed Genet 2002;119:57–63.

15.

Shastry

BS.

SNP alleles in human disease and evolution. J Hum Genet 2002;47:561–566.

16.

Shen

, et al. High-throughput SNP genotyping on universal bead arrays. Mutat Res 2005;573:70–82.

17.

Shi

, et al. Development and application of genotyping technologies. Sci China Ser C Life Sci 2009;52:17–23.

18.

Shimada

, et al. Polynucleotide kinase-phosphatase enables neurogenesis via multiple DNA repair pathways to maintain genome stability. EMBO J 2015;34:2465–2480.

19.

Syvänen A-C. Toward genome-wide SNP genotyping. Nat Genet 2005;37(Suppl):S5–S10.

20.

Tinh

, et al. Polymorphisms of candidate genes associated with growth and carcass traits in Canadian Duroc pigs. Ind J Anim Res 2023;57:831–835.

21.

Walls

, et al. Defining “normal” in pig parturition. Animals (Basel) 2022;12:2754.

22.

Wang

, et al. Ribosome assembly factor URB1 contributes to colorectal cancer proliferation through transcriptional activation of ATF4. Cancer Sci 2020;112:101–116.

23.

Wang

W-P

, et al. [Approaches for SNP genotyping]. Yi Chuan 2006;28:117–126. Chinese.

24.

Yang

, et al. POLR1B is upregulated and promotes cell proliferation in non-small cell lung cancer. Oncol Lett 2020;19:671–680.