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United States Department of Agriculture

Agricultural Research Service

Title: Use of a Multi-Probe Ribonuclease Protection Assay to Detect the Presence of Mrnas of Various Cytokines in Immune and Endocrine System Tissues of Domestic Livestock

Authors
item Abbey, C - TEXAS A&M UNIV.
item Livingston, K - TEXAS A&M UNIV.
item Parent, S - TEXAS A&M UNIV.
item Carroll, Jeffery
item Forbes, T - TEXAS AGRIC. EXP. STN.
item Laurenz, J - TEXAS A&M UNIV.
item Randel, R - TEXAS AGRIC. EXP. STN.
item Welsh, T - TEXAS A&M UNIV.

Submitted to: American Society of Animal Science Southern Section Meeting
Publication Type: Abstract Only
Publication Acceptance Date: February 2, 2000
Publication Date: N/A

Technical Abstract: A multi-probe ribonuclease protection assay system was used to simultaneously detect the presence of mRNAs coding for 8 different cytokines in porcine tissues (spleen, thymus, liver, hypothalamus, pituitary, adrenal, corpus luteum, testis). Tissues were collected from pigs, cattle and goats and frozen at -80 degrees C. Tissues were homogenized and RNA extracted with Tri-Pure Reagent (Boehringer). Ten micrograms of total RNA were hybridized overnight with a panel of radiolabelled porcine cytokine probes [interleukin-1 alpha (IL-1a), IL-1 beta (IL-1b), IL-6, IL-10, IL-12p35, IL-12p40, IL-18, and interferon-gamma (INF-g): PharMingen, San Diego], RNAsed to remove single-stranded nucleic acids, run on an acrylamide gel and exposed overnight on X-ray film. Expression of mRNAs corresponding to IL-6, IL-12p35 and INF-g was not observed. Signals corresponding to IL-1a mRNA and IL-10 mRNA were detected din the spleen and thymus: IL-12p40 mRNA was detected in the thymus. Expression of IL-18 mRNA varied from a moderate level in the testis, thymus, and spleen to a lower level in adrenal and liver; IL-18 mRNA was not observed in the hypothalamus, pituitary and corpus luteum. IL-1b mRNA was detected in the aforementioned tissues at a low level. This porcine specific cytokine panel was used with RNA from goat spleen and liver, and cattle liver. Low levels of IL-18 mRNA and IL-1b mRNA were detected in these tissues. This method can facilitate detection of cytokines associated with reproductive, growth, stress and immune responses.

Last Modified: 7/24/2014