禽腺病毒血清4型感染鸡组织中NLRP3基因转录水平的实时荧光定量PCR分析

doi:10.11843/j.issn.0366-6964.2021.020

Abstract

Abstract: To analyze the transcription level of NLRP3 gene in chickens infected with FAdV-4, a pair of NLRP3 specific primers was designed, and the 180 bp NLRP3 gene was amplified by RT-PCR and cloned into the pMD-18T vector to create a recombinant plasmid pMD-18T-NLRP3. Using the constructed pMD-18T-NLRP3 plasmid as standards, a standard curve assay for SYBR Green I real-time PCR was established. After optimization of reaction conditions, a real-time fluorescent quantitative PCR was successfully established to detect NLRP3, and it was applied to measure the transcription level of NLRP3 gene in the organs of chickens infected with pathogenic FAdV-4. The results indicated that the primers designed for this assay were specific to chicken NLRP3 gene; the established assay was able to provide a well-defined quantification standard curve for the chicken NLRP3 standard plasmid, and a linear correlation was observed between the copy numbers of the standard plasmid pMD-18T-NLRP3 and the Cq values. Comparing to the control group, the transcription level of NLRP3 was significantly higher in the liver and spleen (P<0.001) and higher in the cecal tonsils and bursa of Fabricius (P<0.01) of the infected chickens. The chicken NLRP3 SYBR Green I real-time PCR established in this study was able to detect the transcription level of NLRP3 in different organs of chickens infected with FAdV-4, and the inflammation induced by FAdV-4 infection is highly related to NLRP3.

Key words: fowl adenovirus serotype 4, chicken NLRP3 gene, real-time fluorescence quantitative PCR, tissue inflammation, transcription level

CLC Number:

S855.3

GUO Huifang, LI Ning, WANG Baiyu, QIAO Qilong, HUANG Qing, LI Yongtao, WANG Zeng, ZHAO Jun. Analysis of the Transcription Level of NLRP3 Gene in Fowl Adenovirus Serotype 4 Infected Chicken Tissues by a Real-time Fluorescence Quantitative PCR[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(1): 195-201.

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Analysis of the Transcription Level of NLRP3 Gene in Fowl Adenovirus Serotype 4 Infected Chicken Tissues by a Real-time Fluorescence Quantitative PCR

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