Next Article in Journal
Ethical and Societal Issues Occasioned by Xenotransplantation
Previous Article in Journal
Goat Milk Nutritional Quality Software-Automatized Individual Curve Model Fitting, Shape Parameters Calculation and Bayesian Flexibility Criteria Comparison
Communication

MHC Genotyping by SSCP and Amplicon-Based NGS Approach in Chamois

1
Faculty of Agriculture, University of Zagreb, 10000 Zagreb, Croatia
2
Faculty of Mathematics, Natural Sciences and Information Technologies, University of Primorska, 6000 Koper, Slovenia
3
Environmental Protection College, 3320 Velenje, Slovenia
4
Institute of Vertebrate Biology of the Czech Academy of Sciences, 60365 Brno, Czech Republic
*
Author to whom correspondence should be addressed.
Animals 2020, 10(9), 1694; https://doi.org/10.3390/ani10091694
Received: 10 August 2020 / Revised: 13 September 2020 / Accepted: 16 September 2020 / Published: 18 September 2020
(This article belongs to the Section Animal Genetics and Genomics)
The major histocompatibility complex is a family of genes of central importance in vertebrate adaptive immunity. Genes of the major histocompatibility complex are among the most polymorphic genes ever described in vertebrates, with their number differing greatly among species. Due to their functional significance and exceptional diversity, they represent excellent markers in evolutionary ecology and conservation. Traditional methods were commonly used for their genotyping, but the introduction of next-generation sequencing facilitated more accurate and reproducible genotyping of such polymorphic gene families. Nevertheless, due to the high polymorphism of the major histocompatibility complex genes, genotyping is inherently difficult. Here, we compared the performance of traditional and next-generation sequencing on genotyping of the major histocompatibility complex genes in chamois. Although the major histocompatibility complex system in chamois is quite simple, we found genotyping discrepancies between the two methods in 25% of individuals. Our results show that next-generation sequencing has a higher detection capacity and thus allows for more accurate genotyping of highly polymorphic genes.
Genes of the major histocompatibility complex (MHC) code for cell surface proteins essential for adaptive immunity. They show the most outstanding genetic diversity in vertebrates, which has been connected with various fitness traits and thus with the long-term persistence of populations. In this study, polymorphism of the MHC class II DRB locus was investigated in chamois with Single-Strand Conformation Polymorphism (SSCP)/Sanger genotyping and Ion Torrent S5 next-generation sequencing (NGS). From eight identified DRB variants in 28 individuals, five had already been described, and three were new, undescribed alleles. With conventional SSCP/Sanger sequencing, we were able to detect seven alleles, all of which were also detected with NGS. We found inconsistencies in the individual genotypes between the two methods, which were mainly caused by allelic dropout in the SSCP/Sanger method. Six out of 28 individuals were falsely classified as homozygous with SSCP/Sanger analysis. Overall, 25% of the individuals were identified as genotyping discrepancies between the two methods. Our results show that NGS technologies are better performing in sequencing highly variable regions such as the MHC, and they also have a higher detection capacity, thus allowing a more accurate description of the genetic composition, which is crucial for evolutionary and population genetic studies. View Full-Text
Keywords: major histocompatibility complex; next-generation sequencing; Ion Torrent; Rupicapra rupicapra major histocompatibility complex; next-generation sequencing; Ion Torrent; Rupicapra rupicapra
Show Figures

Figure 1

MDPI and ACS Style

Stipoljev, S.; Bužan, E.; Rolečková, B.; Iacolina, L.; Šprem, N. MHC Genotyping by SSCP and Amplicon-Based NGS Approach in Chamois. Animals 2020, 10, 1694. https://doi.org/10.3390/ani10091694

AMA Style

Stipoljev S, Bužan E, Rolečková B, Iacolina L, Šprem N. MHC Genotyping by SSCP and Amplicon-Based NGS Approach in Chamois. Animals. 2020; 10(9):1694. https://doi.org/10.3390/ani10091694

Chicago/Turabian Style

Stipoljev, Sunčica, Elena Bužan, Barbora Rolečková, Laura Iacolina, and Nikica Šprem. 2020. "MHC Genotyping by SSCP and Amplicon-Based NGS Approach in Chamois" Animals 10, no. 9: 1694. https://doi.org/10.3390/ani10091694

Find Other Styles
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Back to TopTop