Applicability of SSCP analysis for MHC genotyping: fingerprinting of <i>Ovar‐DRB1</i> exon 2 alleles from Finnish and Russian breeds

Kostia, Silja; Kantanen, Juha; Kolkkala, Mikko; Varvio, Sirkka-Liisa

doi:10.1046/j.1365-2052.1998.296376.x

Cited by 19 publications

(18 citation statements)

References 2 publications

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“…The results revealed extensive polymorphism at this locus. In Finnish and Russian breeds, by using the SSCP technique, 7 new sequences and 19 alleles in exon 2 of 31 animals were detected (14). In the Latxa breed, 9 alleles were identified.…”

Section: Discussionmentioning

confidence: 99%

“…Several polymerase chain reaction (PCR)-based methods have been used for studying the polymorphism in MHC in sheep, including sequence-specific oligonucleotide probe analysis (13), single-strand conformation polymorphism (SSCP) (14,15), cloning and sequencing (16), PCR-restriction fragment length polymorphism (RFLP) (11,17), and direct sequence analysis of PCR products (18). To date, more than 160 Ovar-DRB1 alleles have been identified by DNA sequencing of exon 2 from various sheep breeds (19).…”

Section: Introductionmentioning

confidence: 99%

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Association of molecular polymorphism in exon 2 of Ovar-DRB1 gene with weight traits in Iranian Makuie sheep breed

ASHRAFI¹,

Mardani²,

HASHEMI³

et al. 2014

Turk J Vet Anim Sci

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Association of molecular polymorphism in exon 2 of Ovar-DRB1 gene with weight traits in Iranian Makuie sheep breed

ASHRAFI¹,

Mardani²,

HASHEMI³

et al. 2014

Turk J Vet Anim Sci

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“…These genes are usually characterized by high levels of diversity among vertebrates (Garrigan and Hedrick 2003). Because of the high polymorphism of the MHC DRB region, routine typing is generally performed using different techniques, e.g., sequence specific oligonucleotide typing (Schwaiger et al 1993), single strand conformational polymorphism (Kostia et al 1998;Jugo and Vicario 2000), PCR followed by restriction fragment length polymorphism (Dutia et al 1994;Amills et al 1996), and 454 pyrosequencing (Babik et al 2009). In addition, DRB cloning and subsequent sequencing often are inevitable (Nagaoka et al 1999).…”

Section: Introductionmentioning

confidence: 99%

Microsatellite-based genotyping of MHC class II DRB1 gene in Iberian and Alpine ibex

et al. 2011

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In an analysis of a microsatellite locus (OLADRB1) linked to the MHC DRB1 gene of Iberian and Alpine ibex (Capra pyrenaica and Carpa ibex), we detected strong linkage disequilibrium between both loci. The allele length polymorphism at OLADRB1 was unambiguously linked to a particular DRB1 allele. This allowed us to develop a DRB-STR matching method for both ibex species. Validation of the DRB-STR matching method was performed in 160 Iberian ibex from Spain and 98 Alpine ibex from Switzerland and Italy. This simple and relatively inexpensive protocol may find wide applications in a variety of research areas (e.g., mate choice, pathogen-driven selection) and in the biological conservation and management of the Western European ibex populations.

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“…Heterozygosity was significantly higher (96%; P < 0.05, t-test) among the Valachian sheep as described in Table1. Different SSCP patterns of DRB1 gene have been reported earlier (Kostia et al 1998;Jugo and Vicario 2000;Jugo and Vicario 2001). The frequency of the k genogroup was the highest (28%) followed by genogroup e (21%).…”

Section: Discussionmentioning

confidence: 89%

“…Moreover two distinct bands (in case of heterozygotes) and single band (in case of homozygote) appeared in asymmetrical PCR-SSCP can be dissected, purified and sequenced for pin-point sequenced based identification of alleles. Kostia et al (1998) also reported the simplicity and speedy nature of SSCP method to accelerate DRB1 genotyping of sheep breeds. a, c, d, e, g, h, k, m, n, o, p, q, r, t, u, v, w, x, y 76 % In short, the application of asymmetric PCR-SSCP technique can facilitates the good molecular marker system for gene polymorphism detection in a large number of individuals, which is usually difficult by using traditional typing techniques.…”

Section: Discussionmentioning

confidence: 99%

Asymmetric PCR-SSCP: a Useful Tool for Detection of OLA-DRB1(MHC Class Ii) Gene Polymorphism in Slovak Improved Valachian Sheep

Tkáčiková¹,

Bhide²,

Mikula³

2005

Acta Vet. Brno

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Tkáãiková ª., M. R., Bhide I. Mikula: Asymmetric PCR-SSCP: a Useful Tool for Detection of OLA-DRB1(MHC class II) Gene Polymorphism in Slovak Improved Valachian Sheep. Acta Vet. Brno 2005, 74: 275-278.Detection of OLA-DBR1 (exon 2) gene polymorphism is presented in the paper. Rapid and inexpensive polymorphism detection method, namely, single stranded conformation polymorphism (SSCP) was assessed. Modification of the SSCP to asymmetric PCR-SSCP enabled a more simplified assay for the clustering the individuals into distinct groups (profiles) on the basis of band patterns, as only single stranded amplicons were detected. A total of 400 Valachian sheep were included in the study. In this cohort, 25 distinct clusters were noticed. Among 25 groups the frequency of k profile was the highest (28%), followed by profile e (21%), p (16 %), w (9%) and d (9%). The homologous SSCP as well as asymmetric-PCR-SSCP patterns were observed among the twins: This finding has shown the sensitivity of both methods to segregate the individuals on the basis of their allelic forms. MHC, OLA-DRB1 gene, SSCP, polymorphism, sheepThe major histocompatibility complex (MHC) plays a central role in the immune response of vertebrates. The extreme polymorphism in MHC genes enables the host to recognize enormous numbers of foreign peptides to trigger an immune reaction. The class II gene region of the sheep MHC (OLA) has an organization similar to that of humans (Scott et al. 1987). Within this region, two sub-regions, namely, DR and DQ exhibit higher polymorphism (Amills et al. 1998). Among OLA class II genes, the DRB1 locus is highly polymorphic. In particular the polymorphism is present in exon 2, which encodes the antigen-binding site. To date nearly 160 OLA-DRB1 alleles have been recorded from various sheep breeds (Konnai et al. 2003).In the population genetics, analysis of large number of samples is a prerequisite. Though the DNA sequencing is gold standard for most of the phylogenetic studies, the cost requisite for such analysis is quite high as well as it is time consuming. Recently, simple and rapid techniques like PCR-RFLP (restriction fragment length polymorphism) and DGGE (denaturing gradient gel electrophoresis) have been used by different researchers for the detection of MHC gene polymorphism (Aldridge et al. 1998;Konnai et al. 2003). These techniques enable to group the individuals into clusters on the basis of gene polymorphism. However, the amount and cost of restriction enzymes required for analyzing large numbers of samples may jeopardize the use of RFLP.SSCP (single stranded conformation polymorphism) offers a simple and inexpensive method for genotyping. Hitherto, SSCP has been extensively used in biomedical research, especially for rapid bacterial genotyping. In this study we used simple SSCP as well as modified asymmetric PCR-SSCP for DRB1 genotyping in autochthonous Valachian sheep breed. To our knowledge, no report is as yet available elaborating the OLA-DBR1 alleles present in the Valachian breed. Considering the...

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Applicability of SSCP analysis for MHC genotyping: fingerprinting of Ovar‐DRB1 exon 2 alleles from Finnish and Russian breeds

Cited by 19 publications

References 2 publications

Association of molecular polymorphism in exon 2 of Ovar-DRB1 gene with weight traits in Iranian Makuie sheep breed

Association of molecular polymorphism in exon 2 of Ovar-DRB1 gene with weight traits in Iranian Makuie sheep breed

Microsatellite-based genotyping of MHC class II DRB1 gene in Iberian and Alpine ibex

Asymmetric PCR-SSCP: a Useful Tool for Detection of OLA-DRB1(MHC Class Ii) Gene Polymorphism in Slovak Improved Valachian Sheep

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