Development and evaluation of a real-time PCR panel for the detection of 20 immune markers in cattle and sheep.

Veterinary Immunology and Immunopathology
Elena L SassuViskam Wijewardana

Abstract

The establishment of a panel of immune markers is of paramount importance to understand the different transcription patterns of infectious diseases in livestock. The array of commercially available immunological assays for cattle and sheep is currently limited, due to the lack of antibodies for these species. Even though SYBR Green based real time quantitative PCR (qPCR) is the most commonly used method to study cytokine transcription in ruminants, a lack of standardization impairs its implementation in the study of different ruminant diseases. In order to obtain reliable qPCR results, several variables need to be considered: choice of reference genes for optimal normalization, variation of annealing temperature among primer sets, and assay specificity and sensitivity. In this study, we developed and validated a panel of immune markers in bovine and ovine samples using SYBR Green based qPCR in a cost-effective way with multiple primer sets optimised to amplify at a common thermal cycling temperature. Twenty primer sets were designed to quantify immune markers (IL-1b, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, IL-13, IL-15, IL-18, IL-23, TNF-α, IFN-γ, IFN-α, Ki-67, NFkB-65, TLR-3, TLR-4, TLR-8 and Rig-1) in ovine and bovine templates...Continue Reading

Citations

Jul 3, 2021·Frontiers in Immunology·Tesfaye Rufael ChibssaCharles Euloge Lamien

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