关键词: 细胞因子, 模式识别受体, 基因表达, 大蒲莲, 长白, 仔猪

Abstract: 【Objective】 Though Chinese indigenous breeds have stronger disease resistance and immunity than imported commercial breeds, little is known about the genetic basis and immune mechanism of their difference. It is supposed that the gene expression level of cytokines and receptors may be the reasons leading to the different disease resistances among breeds. In the present study, the mRNA expression levels of seven genes in piglets of Dapulian and Landrace were determined, including two pro-inflammatory cytokines (IL8 and TNFα), one anti-inflammatory cytokine (IL10), one interferon regulatory factor (IRF3), two interferon (IFNA and IFNG) and one pattern recognition receptor (TLR4), and their breed and sow effects in the two breeds were analyzed, which would benefit the understanding of the immunity levels of Chinese indigenous breeds and the commercial breeds. 【Method】 Blood samples of clinically healthy Dapulian (n=104) and Landrace (n=171) piglets of 35 days old were collected, and their total RNA was extracted and reverse-transcribed into cDNA. Based on the quantitative real-time PCR, firstly, the expression stabilities of the six candidate genes (ACTB, GAPDH, B2M, TBP, RPL4 and PPIA) were evaluated by the geNorm program to select the most stable reference gene. Then, the selected reference gene was used for normalization, and the gene expression levels of the above mentioned cytokine and receptor genes were quantified in piglets of Dapulain and Landrace. Finally, the breed and sow effects of their expression levels in the two breeds were analyzed by Least Square Means. 【Result】 The expression stability values (M) of the six candidate reference genes in blood were as follows: ACTB (0.595) ＞ GAPDH＞ PPIA＞ RPL4＞ TBP＞ B2MΔCt of the six cytokine genes were 9.23-14.25 and 5.83-8.73, respectively, indicating there were large difference among individuals of the two tested breeds. Compared with six cytokines, the D-values of TLR4 were smaller, 5.03 and 3.82 for Dapulian and Landrace respectively. Comparison between the two breeds, the expression levels of the six cytokines in Landrace were higher than that of Dapulian, while the expression levels of TLR4 in Dapulian was higher than that of Landrace. Additionally, the variation coefficients of all the cytokine and receptor genes in Dapulian (the average is 31.43%) were larger than those in Landrace (the average is 19.10%), which is consistent with the fact that Dapulian is one indigenous breed with less selecting strength compared with Landrace. The breed effect analyses indicated that, except IL10＜0.05) level or 0.01 level (P＜0.01). The sow effect analysis showed that it had significant influence on all the genes determined at 0.01 level (P＜0.01). 【Conclusion】 In the present study, the stably expressed reference genes were successfully selected which are used for the gene expression research of porcine peripheral blood, seven genes’ mRNA expression levels of peripheral blood in normally raised Dapulian and Landrace piglets were determined at population level and the breed and sow effects on their expression were analyzed. Results of the study would provide important information concerning understanding of the variation between the indigenous and commercial breeds and mining the characteristics of indigenous breed. , breed effect had significant influence on all the other genes determined at 0.05 (P (0.440). B2M, which had the most stable expression, was selected as the reference gene. In Dapulian and Landrace, the D-values between the maximum and minimum of (0.461) (0.466) (0.469) (0.571)

Key words: cytokines, pattern recognition receptors, gene expression, Dapulian, Landrace, piglet