A multiplex PCR genotyping assay to distinguish XX and XY tissues in sheep
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A multiplex PCR genotyping assay to distinguish XX and XY tissues in sheep River Evans1 · Heidi J. Engelhorn1,2 · Quinton A. Winger1 · Adam J. Chicco1 · Gerrit J. Bouma1 Received: 17 June 2020 / Revised: 21 July 2020 / Accepted: 26 July 2020 © Springer Nature B.V. 2020
Abstract Sex is an important biological variable as many physiological as well as disease processes differ between females and males. The fundamental biological distinction between females and males starts with chromosomal sex, and the establishment of XX and XY cells and tissues. Polymerase Chain Reaction (PCR) is a simple and effective method to easily determine chromosomal or genetic sex of cells and tissues. The goal of this study was to develop a simple multiplex PCR genotyping assay to distinguish XX and XY tissues in sheep. Primers were designed to amplify a fragment of the autosomal gene myogenin (MYOG) and sex determining region on the Y chromosome (SRY). PCR analysis was performed on a variety of genomic DNA samples isolated from fetal sheep skeletal muscle, brain, liver, and placenta, and revealed a single 259 bp band for MYOG in XX females, and a 259 bp band for MYOG and a 167 bp band for SRY in XY males. Amplicons were clearly distinguishable by gel electrophoresis, and their sequences revealed 100% identity to the known ovine MYOG and SRY sequence. The reported multiplex PCR genotyping assay provides a rapid means to distinguish XX and XY sheep tissues using low volume samples. Keywords XX · XY · Sheep · PCR · Genotyping Abbreviations PCR Polymerase chain reaction MYOG Myogenin SRY Sex determining region on the Y chromosome AMEL Amelogenin
River Evans and Heidi J. Engelhorn contributed equally to this work. * Gerrit J. Bouma [email protected] River Evans [email protected] Heidi J. Engelhorn [email protected] Quinton A. Winger [email protected] Adam J. Chicco [email protected] 1
Department of Biomedical Sciences, Animal Reproduction & Biotechnology Laboratory, Colorado State University, Fort Collins, CO 80523, USA
Colorado Center for Reproductive Medicine, 10290 RidgeGate Circle, Lone Tree, CO 80124, USA
2
Background For decades, mouse models have been used extensively in research studies due to the ability for genome modifications, large litter sizes, and short gestation time. However, physiological and physical differences between mice and humans have limited research progress in some areas, and other potentially more relevant animal models such as the sheep have been used. For example, similar to humans, sheep have a longer gestation period, generally give birth to single or twin offspring, and have shown close similarities in fetal development and pregnancy disorders such as intrauterine growth restriction [1]. An important fundamental aspect of physiology are the known differences between females and males, which either directly or indirectly can be attributable to the sex (X and Y) chromosomes. In biomedical research using animal models it is importa
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