Skip to main content
SpringerLink
Log in

Rapid assignment of swine leukocyte antigen haplotypes in pedigreed herds using a polymerase chain reaction-based assay

  • Original Paper
  • Published:
Immunogenetics Aims and scope Submit manuscript

Abstract

We present a simple assay to determine the swine leukocyte antigen (SLA) haplotypes of animals within two experimental populations of MHC defined miniature pigs. The Yucatan miniature pigs have four founder haplotypes (w, x, y, z) and one recombinant haplotype (q). The NIH miniature pigs have three founder haplotypes (a, c, d) and two recombinant haplotypes (f, g). Because most crossovers occur between the class I and class II regions, haplotypes can be assigned by typing one class I locus and one class II locus for practical purposes. We have previously characterized these seven founder haplotypes by sequencing the cDNA of three SLA class I loci, designated as SLA-1, SLA-3 and SLA-2 and four SLA class II loci, SLA-DQA1, SLA-DQB1, SLA-DRA1 and SLA-DRB1. These sequences were used to design allele-specific primers to amplify one MHC class I and one MHC class II gene for each haplotype. Primers were tested for specificity in homozygous and heterozygous animals. Positive control primers were also designed to amplify a portion of the E-selectin or α-actin gene and multiplexed with the allele-specific primers to check for false negatives. This combination of allele-specific and positive control primers produced specific and robust PCR-site-specific primer assays for assigning SLA haplotypes in the two populations.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1a, b.
Fig. 2.

Similar content being viewed by others

References

  • Akane A, Matsubara K, Nakamura H, Takahashi S, Kimura K (1994) Identification of the heme compound copurified with deoxyribonucleic acid (DNA) from bloodstains, a major inhibitor of polymerase chain reaction (PCR) amplification. J Forensic Sci 39:362–372

    CAS  PubMed  Google Scholar 

  • Butler JE, Lunney JK (1998) Immunogenetics. In: Rothschild MF, Ruvinsky A (ed) Genetics of the pig. CAB International, Wallingford, UK, pp 163–197

  • Frels WI, Bordallo C, Golding H, Rosenberg A, Rudikoff S, Singer DS (1990) Expression of a class I MHC transgene: regulation by a tissue-specific negative regulatory DNA sequence element. New Biologist 2:1024–1033

    CAS  PubMed  Google Scholar 

  • Gustafsson K, Germana S, Hirsch F, Pratt K, LeGuern C, Sachs DH (1990a) Structure of miniature swine class II DRB genes: conservation of hypervariable amino acid residues between distantly related mammalian species. Proc Natl Acad Sci USA 87:9798–9802

    CAS  PubMed  Google Scholar 

  • Gustafsson K, LeGuern C, Hirsch F, Germana S, Pratt K, Sachs DH (1990b) Class II genes of miniature swine. IV. Characterization and expression of two allelic class II DQB cDNA clones. J Immunol 145:1946–1951

    CAS  PubMed  Google Scholar 

  • Hoppe BL, Conti-Tronconi BM, Horton RM (1992) Gel-loading dyes compatible with PCR. Biotechniques 12:679–680

    CAS  PubMed  Google Scholar 

  • Hosokawa-Kanai T, Tanioka Y, Tanigawa M, Matsumoto Y, Ueda S, Onodera T (2002) Differential alloreactivity at SLA-DR and -DQ matching in two-way mixed lymphocyte culture. Vet Immunol Immunopathol 85:77–84

    Article  CAS  PubMed  Google Scholar 

  • Ierino FL, Gojo S, Banerjee PT, Giovino M, Xu Y, Gere J, Kaynor C, Awwad M, Monroy R, Rembert J, Hatch T, Foley A, Kozlowski T, Yamada K, Neethling FA, Fishman J, Bailin M, Spitzer TR, Cooper DK, Cosimi AB, LeGuern C, Sachs DH (1999) Transfer of swine major histocompatibility complex class II genes into autologous bone marrow cells of baboons for the induction of tolerance across xenogeneic barriers. Transplantation 67:1119–1128

    CAS  PubMed  Google Scholar 

  • Lumsden JS, Kennedy BW, Mallard BA, Wilkie BN (1993) The influence of the swine major histocompatibility genes on antibody and cell-mediated immune responses to immunization with an aromatic-dependent mutant of Salmonella typhimurium. Can J Vet Res 57:14–18

    CAS  PubMed  Google Scholar 

  • Lunney JK (1994) Current status of the swine leukocyte antigen complex. Vet Immunol Immunopathol 43:19–28

    Article  CAS  PubMed  Google Scholar 

  • Mallard BA, Kennedy BW, Wilkie BN (1991) The effect of swine leukocyte antigen haplotype on birth and weaning weights in miniature pigs and the role of statistical analysis in this estimation. J Anim Sci 69:559–564

    CAS  PubMed  Google Scholar 

  • Martin M, Mann D, Carrington M (1995) Recombination rates across the HLA complex: use of microsatellites as a rapid screen for recombinant chromosomes. Hum Mol Gen 4:423–428

    PubMed  Google Scholar 

  • Mytilineos J, Lempert M, Middleton D, Williams F, Cullen C, Scherer S, Opelz G (1997) HLA class I DNA typing of 215 "HLA-A, -B, -DR zero mismatched" kidney transplants. Tissue Antigens 50:355–358

    CAS  PubMed  Google Scholar 

  • Noreen HJ, Yu N, Setterholm M, Ohashi M, Baisch J, Endres R, Fernandez-Vina M, Heine U, Hsu S, Kamoun M, Mitsuishi Y, Monos D, Perlee L, Rodriguez-Marino S, Smith A, Yang SY, Shipp K, Hegland J, Hurley CK (2001) Validation of DNA-based HLA-A and HLA-B testing of volunteers for a bone marrow registry through parallel testing with serology. Tissue Antigens 57:221–229

    Article  CAS  PubMed  Google Scholar 

  • Olerup O, Aldener A, Fogdell A (1993) HLA-DQB1 and -DQA1 typing by PCR amplification with sequence-specific primers (PCR-SSP) in 2 hours. Tissue Antigens 41:119–134

    CAS  PubMed  Google Scholar 

  • Pennington LR, Lunney JK, Sachs DH (1981) Transplantation in miniature swine. VIII. Recombination within the major histocompatibility complex of miniature swine. Transplantation 31:66–71

    CAS  PubMed  Google Scholar 

  • Rychlik W, Spencer WJ, Rhoads RE (1990) Optimization of the annealing temperature for DNA amplification in vitro. Nucleic Acids Res 18:6409–6412

    PubMed  Google Scholar 

  • Sachs DH, Leight G, Cone J, Schwarz S, Stuart L, Rosenberg S (1976) Transplantation in miniature swine. I. Fixation of the major histocompatibility complex. Transplantation 22:559–567

    CAS  PubMed  Google Scholar 

  • Satz ML, Wang LC, Singer DS, Rudikoff S (1985) Structure and expression of two porcine genomic clones encoding class I MHC antigens. J Immunol 135:2167–2175

    CAS  PubMed  Google Scholar 

  • Schook LB, Rutherford MS, Lee JK, Shia YC, Bradshaw M, Lunney JK (1996) The swine major histocompatibility complex. In: Schook LB, Lamont SJ (eds) The major histocompatibility complex of domestic animal species. CRC Press, New York, pp 212–214

  • Scott I, O'Shea J, Bunce M, Tiercy JM, Arguello JR, Firman H, Goldman J, Prentice HG, Little AM, Madrigal JA (1998) Molecular typing shows a high level of HLA class I incompatibility in serologically well matched donor/patient pairs: implications for unrelated bone marrow donor selection. Blood 92:4864–4871

    CAS  Google Scholar 

  • Shia YC, Bradshaw M, Rutherford MS, Lewin HA, Schook LB (1995) Polymerase chain reaction based genotyping for characterization of SLA-DQB and SLA-DRB alleles in domestic pigs. Anim Genet 26:91–100

    CAS  PubMed  Google Scholar 

  • Singer DS, Ehrlich R, Satz L, Frels W, Bluestone J, Hodes R, Rudikoff S (1987) Structure and expression of class I MHC genes in the miniature swine. Vet Immunol Immunopathol 17:211–221

    Article  CAS  PubMed  Google Scholar 

  • Sullivan JA, Oettinger HF, Sachs DH, Edge AS (1997) Analysis of polymorphism in porcine MHC class I genes: alterations in signals recognized by human cytotoxic lymphocytes. J Immunol 159:2318–2326

    CAS  PubMed  Google Scholar 

  • Termijtelen A, van Rood JJ (1981) Complexity of stimulation in MLC and the influence of matching for HLA-A and -B. Scand J Immunol 14:459–466

    CAS  PubMed  Google Scholar 

  • Thistlethwaite JR Jr, Pennington LR, Lunney JK, Sachs DH (1983) Immunologic characterization of MHC recombinant swine. Production of SLA class-specific antisera and detection of Ia antigens on both B and non-B PBL. Transplantation 35:394–400

    PubMed  Google Scholar 

  • Tissot RG, Beattie CW, Amoss MS Jr (1989) The swine leucocyte antigen (SLA) complex and Sinclair swine cutaneous malignant melanoma. Anim Genet 20:51–57

    CAS  PubMed  Google Scholar 

  • Vaiman M, Chardon P, Renard C (1979) Genetic organization of the pig SLA complex. studies on nine recombinants and biochemical and lysostrip analysis. Immunogenetics 9:356–361

    Google Scholar 

  • Velten F, Rogel-Gaillard C, Renard C, Pontarotti P, Tazi-Ahnini R, Vaiman M, Chardon P (1998) A first map of the porcine major histocompatibility complex class I region. Tissue Antigens 51:183–194

    CAS  PubMed  Google Scholar 

  • Voorter CE, Kik MC, van den Berg-Loonen EM (1998) High-resolution HLA typing for the DQB1 gene by sequence-based typing. Tissue Antigens 51:80–87

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors wish to thank Dr. Guy Bouchard of the Sinclair Research Center, Columbia, Mo. for providing samples for the Yucatan Miniature pigs and Dr. Lyn Colenda of the NIH Animal Center, Beltsville, Md. for providing some of the samples for the NIH Miniature pigs.

Author information

Authors and Affiliations

  1. Pathology Department, Transplant Immunology Lab, Baylor University Medical Center, 3500 Junius St., Dallas, TX 75246, USA

    Gregory W. Martens, Judy E. Baker & Douglas M. Smith

  2. Immunology and Disease Resistance Laboratory, ANRI, ARS, USDA, Beltsville, MD 20705-2350, USA

    Joan K. Lunney

Authors
  1. Gregory W. Martens
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Joan K. Lunney
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Judy E. Baker
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Douglas M. Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Douglas M. Smith.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Martens, G.W., Lunney, J.K., Baker, J.E. et al. Rapid assignment of swine leukocyte antigen haplotypes in pedigreed herds using a polymerase chain reaction-based assay. Immunogenetics 55, 395–401 (2003). https://doi.org/10.1007/s00251-003-0596-3

Download citation

  • Received:

  • Revised:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00251-003-0596-3

Keywords

Search

Navigation