The impact of porcine reproductive and respiratory syndrome virus infection on lung cytokine gene expression by reverse transcription-quantitative competitive polymerase chain reaction

• Main Authors: Yen, Hong-Hsin, 顏宏勳
• Other Authors: Chian-Ren Jeng
• Format: Others
• Language: zh-TW
• Published: 1998
• Online Access: http://ndltd.ncl.edu.tw/handle/44080832747843525720

碩士 === 國立臺灣大學 === 獸醫學系研究所 === 86 === In this study, we used reverse transcription-quantitative competitive pol ymerase chain reaction（RT-qcPCR）to assay porcine cytokines and porcine repro ductive and respiratory syndrome virus（PRRSV）mRNA expression, and investigat ed the effect of PRRSV infection on cytokine gene expression in the lung tissu e of nursing pigs. We modified a previously described RT-qcPCR method to const ruct a DNA fragment to measure interleukin-1beta（IL-1beta）, IL-2, IL-4, IL-8 , interferon-gamma（IFN-gamma）,tumor necrosis factor-alpha, beta-actin, and P RRSV mRNA expression in pig. This constructed DNA fragment had identical prime r sequences to target cDNA except differences in the size of products. After c o-amplifying of RNA-derived cDNA with serial dilution of the DNA fragment and calculating the ratios between them, a precise quantification data of sample w as obtained.By using this DNA fragment, it was possible to analyze IL-1beta, I FN-gamma and PRRSV mRNA expression in the lung between PRRSV-infected（n=4）an d sham-inoculated（n=4）piglets. The results revealed that piglets infected by PRRSV had a higher level of IFN-gamma mRNA expression in their lung than thos e of sham-inoculation（P＜0.05）at 7, 14, and 21 days post-inoculation（PI）. PRRSV-infected piglets also showed higher IL-1beta mRNA expression than sham-i noculated ones at 21 days PI（P＜0.05）although there were no differences betw een the groups of 7 and 21days PI. Besides, as virus load increased IFN-gamma and IL-1beta mRNA expression would also elevated. From these results, it is co ncluded that PRRSV infection would alter IFN-gamma and IL-1beta mRNA levels in the lung tissue of infected piglets; however thedisturbance of immune system in piglet by PRRSV infection required further study.