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Immunogenetics

, Volume 64, Issue 8, pp 633–640 | Cite as

Rapid discrimination of MHC class I and killer cell lectin-like receptor allele variants by high-resolution melt analysis

  • Alyssa Lundgren
  • Sharon Kim
  • Michael D. Stadnisky
  • Michael G. Brown
Brief Communication

Abstract

Ly49G and H-2 class I Dk molecules are critical to natural killer cell-mediated viral control. To examine their contributions in greater depth, we established NK gene complex (NKC)/Ly49 congenic strains and a novel genetic model defined by MHC class I Dk disparity in congenic and transgenic mouse strains. Generation and maintenance of Ly49 and H-2 class I select strains require efficient and reproducible genotyping assays for highly polygenic and polymorphic sequences. Thus, we coupled gene- and allele-specific PCR with high-resolution melt (HRM) analysis to discriminate Ly49g and H-2 class I D and K alleles in select strains and in the F2 and backcross hybrid offspring of different genetic crosses. We show that HRM typing for these critical immune response genes is fast, accurate, and dependable. We further demonstrate that H-2 class I D HRM typing is competent to detect and quantify transgene copy numbers in different mice with distinct genetic backgrounds. Our findings substantiate the utility and practicality of HRM genotyping for highly related genes and alleles, even those belonging to clustered multigene families. Based on these findings, we envision that HRM is capable to interrogate and quantify gene- and allele-specific variations due to differential regulation of gene expression.

Keywords

Genotyping H-2 D Klra7 Ly49g NK gene complex Transgene copy number 

Notes

Acknowledgments

This research was supported with Public Health Service grants (AI050072 and AI082024) from the National Institutes of Health, National Institute of Allergy and Infectious Diseases. We thank Jessica Prince, Jack Cronk, Heather Lee, and Thuan Nguyen for technical support.

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Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Alyssa Lundgren
    • 1
    • 2
  • Sharon Kim
    • 3
  • Michael D. Stadnisky
    • 1
    • 2
    • 4
    • 5
  • Michael G. Brown
    • 1
    • 2
    • 4
  1. 1.Department of Medicine, Division of NephrologyUniversity of Virginia School of MedicineCharlottesvilleUSA
  2. 2.Carter Immunology Center for Immunology ResearchUniversity of Virginia School of MedicineCharlottesvilleUSA
  3. 3.Department of BiologyUniversity of VirginiaCharlottesvilleUSA
  4. 4.Department of Microbiology, Immunology and Cancer BiologyUniversity of Virginia School of MedicineCharlottesvilleUSA
  5. 5.Tree Star, IncAshlandUSA

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