InDel Variations of PRL and GHR Genes Associated with Litter Size in Pirlak sheep breed

Year 2023, Volume: 20 Issue: 4, 890 - 897, 25.12.2023

Sude Atay , Kadriye Gül Yurdagül , Ümit Bilginer , Taki Karslı , Eymen Demir

https://doi.org/10.33462/jotaf.1226643

Abstract

Numerous molecular genotyping methods are available to analyse local livestock populations at molecular level in which traditional Polymerase Chain Reaction (PCR) guided by specific oligonucleotides is a fast and cost-effective method to investigate single genes. Until today, many genes which are of major effects on litter size have been reported in sheep. Genetic variations in these genes shaping the expression profile at DNA level may lead to differences in litter size among the sheep breeds. This is the first attempt to investigate insertion/deletion (inDel) variations in Prolactin (PRL) intron 2 and Growth Hormone Receptor (GHR) intron 3 and intron 4 genes in Pirlak sheep breed via traditional PCR technique. A total of 100 unrelated animals sampled from representative herds reared in Antalya were genotyped based on absence/presence of 23 base pairs (bp) length inDel in which three genotypes (II, ID, and DD) were detected in all loci. I and D allele frequency were 0.421 and 0.579, respectively in terms of PRL-intron 2 locus. I / D allele frequencies were found as 0.599 / 0.401 and 0.372 / 0.628 in GHR intron 3 and intron 4, respectively. The lowest II (0.181) and DD (0.177) genotype frequencies were detected in GHR-intron 4 and GHR-intron 3 loci, respectively. The lowest (0.177 for DD) and highest (0.448 for ID) genotype frequencies were detected in GHR intron 3 locus across the population. Significant deviation from Hardy-Weinberg Equilibrium (HWE) was detected only in PRL-intron 2 locus. The results of the present study confirm that Pirlak breed conserves sufficient genetic variation in PRL and GHR gene regions which could be utilized in selection strategies in order to increase litter size in the future.

Keywords

Deletion, Insertion, Litter size, Multiple birth, PCR

Supporting Institution

Scientific and Technological Research Council of Turkey

Project Number

1919B012101328

Thanks

This work supported by the Scientific and Technological Research Council of Turkey (Project No: 1919B012101328), Republic of Turkey.

Pırlak Koyun Irkında Bir Batında Doğan Yavru Sayısı ile Ilişkili PRL ve BHR Genlerindeki inDel Varyasyonlar

Abstract

Çiftlik hayvanı populasyonlarının moleküler seviyede analiz edilmesine olanak sağlayan çok sayıda moleküler genotipleme yöntemleri bulunmakla birlikte özgün oligonükleotidlerle yönlendirilmiş geleneksel Polimeraz Zincir Reaksiyonu (PZR) tekli genlerin incelenmesinde kullanılan hızlı ve uygun maliyetli bir tekniktir. Günümüze kadar koyunlarda bir batında doğan yavru sayısı üzerine major etkileri olan çok sayıda gen bildirilmiştir. DNA seviyesinde ekspresyon profilini şekillendiren bu genlerdeki varyasyonlar koyun ırklarında bir batında doğan yavru sayısında farklılıkların görülmesine neden olabilmektedir. Bu çalışma Pırlak koyun ırkında Prolaktin (PRL) intron 2 ve Büyüme Hormonu Reseptörü (BHR) intron 3 ve intron 4 gen bölgelerindeki insersiyon/delesyon (inDel) varyasyonlarının geleneksel PZR yöntemiyle incelendiği ilk araştırmadır. Antalya’da yetiştirilen, ırkı temsil eden ve birbiriyle akraba olmayan toplam 100 hayvan 23 baz çifti (bç) uzunluğundaki inDel’in varlığı/yokluğuna göre genotiplendirilmiş ve bütün varyasyonlar bakımından üç farklı genotip (II, ID ve DD) tespit edilmiştir. PRL-intron 2 lokusunda I ve D allel frekansı sırasıyla 0.421 ve 0.579 olarak bulunmuştur. BHR-intron 3 ve intron 4 lokuslarındaki I / D allel frekansları sırasıyla 0.599 / 0.401 ve 0.372 / 0.628 olarak bulunmuştur. En düşük II (0.181) ve DD (0.177) genotip frekansı sırasıyla BHR-intron 4 ve BHR-intron 3 lokusunda belirlenmiştir. Populasyon seviyesinde en düşük (DD için 0.177) ve en yüksek (ID için 0.448) genotip frekasnları BHR intron 3 lokusunda tespit edilmiştir. Sadece PRL-intron 2 lokusunda Hardy-Weinberg (HWD) dengesinden önemli sapma tespit edilmiştir. Bu çalışmanın sonuçları Pırlak ırkının PRL ve BHR gen bölgeleri bakımından yeterli seviyede genetik varyasyona sahip olduğunu ve bunun gelecekte bir batında doğan yavru sayısını arttırmak için yapılacak seleksiyon stratejilerinde kullanılabileceğini göstermiştir.

Keywords

Delesyon, Insersiyon, Bir batında doğan yavru sayısı, Çoklu doğum, PZR

Project Number

1919B012101328

References

• Akhatayeva, Z., Mao, C., Jiang, F., Pan, C., Lin, C., Hao, K., Lan, T., Chen, H., Zhang, Q. and Lan, X. (2020). Indel variants within the PRL and GHR genes associated with sheep litter size. Reproduction in Domestic Animals, 55(11): 1470-1478.
• Al-Anbari, N. N., Azzawi, M. S., Riyadh, H. M., WedadSalih, D. and Raghad, A. H. (2018). Estimation of genetic variance using data of prolactin gene in Turkish Awassi sheep. Journal of Entomology and Zoology Studies, 6(2): 727-729.
• Al-Thuwaini, T. M. (2021). Novel single nucleotide polymorphism in the prolactin gene of Awassi ewes and its role in the reproductive traits. Iraqi Journal of Veterinary Sciences, 35(3): 429-435.
• Anonymous (2023). Turkish Statistical Institute. https://data.tuik.gov.tr/Kategori/GetKategori?p=tarim-111&dil=1 (Accessed date: 23.08.2023).
• Bodin, L., Di Pasquale, E., Fabre, S., Bontoux, M., Monget, P., Persani, L. and Mulsant, P. (2007). A novel mutation in the bone morphogenetic protein 15 gene causing defective protein secretion is associated with both increased ovulation rate and sterility in Lacaune sheep. Endocrinology, 148(1): 393-400.
• Cam, M. A. and Kuran, M. (2004). Effects of a single injection of hCG or GnRH agonist on day 12 post mating on fetal growth and reproductive performance of sheep. Animal Reproduction Science, 80(1-2): 81-90.
• Çelikeloğlu, K., Erdoğan, M., Hacan, Ö., Koçak, S., Bozkurt, Z. and Tekerli, M. (2018). Investigation of possible polymorphisms in BMPR1B, BMP15 and GDF9 genes in Pırlak sheep. Kocatepe Veterinary Journal, 11(4): 356-362.
• Davis, G. H., Montgomery, G. W., Allison A. J., Kelly, R. W. and Bray, A. R. (1982). Segregation of a major gene influencing fecundity in progeny of Booroola sheep. New Zealand Journal of Agricultural Research, 25(4): 525-529.
• Demir, E., Ceccobelli, S., Bilginer, U., Pasquini, M., Attard, G. and Karsli, T. (2022). Conservation and Selection of Genes Related to Environmental Adaptation in Native Small Ruminant Breeds: A Review. Ruminants, 2(2): 255-270.
• Ertuğrul, M., Dellal, G., Soysal, İ., Elmacı, C., Akın, O., Arat, S., Barıtçı, İ., Pehlivan, E. and Yılmaz, O. (2009). Protection of native sheep breeds of Turkey. Journal of Agricultural Faculty of Uludag University, 23(2): 97-119.
• Esmaeili-Fard, S. M., Gholizadeh, M., Hafezian, S. H. and Abdollahi-Arpanahi, R. (2021). Genome-wide association study and pathway analysis identify NTRK2 as a novel candidate gene for litter size in sheep. PloS One 16(1): e0244408.
• GDARP (2009). Introduction catalogue of Domestic Animal Genetic Resources of Türkiye. https://www.tarimorman.gov.tr/TAGEM/Belgeler /yayin/Katalog%20Türkçe.pdf (Accessed date : 11.05.2023).
• Ghiasi, H. and Abdollahi-Arpanahi, R. (2021). The candidate genes and pathways affecting litter size in sheep. Small Ruminant Research, 205: 106546.
• Hanrahan, J. P., Gregan, S. M., Mulsant, P., Mullen, M., Davis, G. H., Powell, R. and Galloway, S. M. (2004).
• Mutations in the genes for oocyte-derived growth factors GDF9 and BMP15 are associated with both increased ovulation rate and sterility in Cambridge and Belclare sheep (Ovis aries). Biology of Reproduction, 70 (4): 900-909.
• Karsli, T. and Balcioglu, M. S. (2010). An investigation of presence of FecB allele on BMPR-IB (Boroola) gene raised in Turkey in six local sheep breeds using PCR-RFLP method. Kafkas Universitesi Veteriner Fakultesi Dergisi, 16(6): 1033-1036.
• Karsli, T., Sahin, E., Karsli, B. A., Eren, M. G. and Balcıoğlu, M. S. (2011). An investigation of presence of FecB, FecXG, FecXH allele in Kangal and Güney Karaman sheep using PCR-RFLP method. Lalahan Hayvancılık Araştırma Enstitüsü Dergisi, 51(2): 71-80.
• Karsli, T., Sahin., Karsli, B. A., Alkan, S. and Balcıoğlu, M. S. (2012). An investigation of mutations (FecXG, FecXI, FecXH, FecXB) on BMP15 gene in some local sheep breeds raised in Turkey. Akdeniz University Journal of the Faculty of Agriculture, 25(1): 29-33.
• Kirikci, K., Cam, M. A. and Mercan, L., (2021). Investigation of G1 (c. 260G> A) polymorphism in exon 1 of GDF9 gene in Turkish sheep breed Karayaka. Turkish Journal of Veterinary and Animal Sciences, 45(1): 191-197.
• Kocaman, İ. and Günal, R. (2007). A research on present structural status improvement sheep barns in the central town of Tekirdag. Journal of Tekirdag Agricultural Faculty, 4 (3), 339-346.
• Koyuncu, M. and Canbolat, O., (2009). Effect of different dietary energy levels on the reproductive performance of Kivircik sheep under a semi-intensive system in the South-Marmara region of Turkey. Journal of Animal and Feed Sciences, 18(4): 620-627.
• Koyuncu, M., Kara Uzun, Ş. and Tuncel, E. (2005). Characterization of goat husbandry and production aspects in South Marmara region. I. General characteristics of goat husbandry. Journal of Agricultural Sciences, 11(4): 373-378.
• Mao, C., Akhatayeva, Z., Cheng, H., Zhang, G., Jiang, F., Meng, X., Yao, Y., Elnour, I. E., Lan, X. and Song, E. (2021). A novel 23 bp indel mutation in PRL gene is associated with growth traits in Luxi Blackhead sheep. Animal Biotechnology, 32(6): 740-747.
• Miller, S., Dykes, D. and Polesky, H. (1988). A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acids Research, 16(3): 1215-1215.
• Narzisi, G. and Schatz, M. C. (2015). The challenge of small-scale repeats for indel discovery. Frontiers in Bioengineering and Biotechnology, 3: 8.
• Öner, Y., Orman, A., Üstüner, H. and Yilmaz, A. (2014). Investigation of polymorphisms on ABCG2, AA-NAT and FABP3 genes in the Kivircik sheep reared in three different provinces of Turkey. Kafkas Universitesi Veteriner Fakultesi Dergisi, 20(5): 649-654.
• Özçelik, U. and Bayram, İ. (2012). The effect of Black Cumin (Nigella sativa) on performance, some blood and rumen fluid parametersin the lambs. Kocatepe Veterinary Journal, 5(2): 27-33.
• Ozmen, O. and Kul, S. (2016). Identification of novel SNPs of ovine PRL gene and their association with milk production traits. Animal Genetics, 52(9): 977-984.
• Ozmen, O., Seker, I., Ertugrul, O., Ozkan, E. and Tekin, N. (2011). Prolactin receptor (PRLR) gene polymorphism in Chios, White Karaman and Awassi sheep breeds. Archives Animal Breeding, 54(4): 381-390.
• Peakall, R. and Smouse, P. E. (2012). GenAlEx 6.5: Genetic analysis in Excel. Population genetic software for teaching and research-an update. Bioinformics, 6(19): 2537-2539.
• Ran, D., Jing, Y., Ming-Xing, C., Gui-Ling, C., Tao, F., Li, F. and Zhong-Xiao, Z. (2011). DNA polymorphism of introns 1 and 2 of prolactin receptor gene and its association with litter size in goats. Animal Science Papers and Reports, 29(4): 343-350.
• Şişman, C. B., Yılmaz, F. and Gezer, E. (2009). Structural situation of sheep barn in Bolu district and improving possibilities. Journal of Tekirdag Agricultural Faculty, 6 (2): 179-189.
• Tao, L., He, X. Y., Wang, F. Y., Pan, L. X., Wang, X. Y., Gan, S. Q. and Chu, M. X. (2021). Identification of genes associated with litter size combining genomic approaches in Luzhong mutton sheep. Animal Genetics, 52(4): 545-549.
• Zhou, M., Pan, Z., Cao, X., Guo, X., He, X., Sun, Q., Di, R., Hu, W., Wang, X., Zhang, X., Zhang, J., Zhang, C., Liu, Q. and Chu, M. (2018). Single nucleotide polymorphisms in the HIRA gene affect litter size in Small Tail Han Sheep. Animals, 8(5): 1-13.