禽流感病毒H9N2亚型三重PCR检测方法的建立

Abstract

Abstract: A triplex reverse transcription-polymerase chain reaction (triplex RT-PCR) was developed to detect avian influenza viruses (AIVs) and H9 N2 subtype AIVs simultaneously. Three pairs of specific primers were designed according to the conserved regions on the sequences of H9 AIV HA gene, N2 AIV NA gene and AIV M gene in GenBank. It showed that all samples containing H9N2 subtype AIV could be amplified into three specific bands, 313 bp for HA gene, 451 bp for NA gene and 667 bp for M gene by this triplex PCR. All samples containing N2 subtype AIV with different HA genes (not H9) could be amplified into two specific bands, 451 bp for N2 subtype AIV and 667 bp for M gene. All samples containing other subtypes of AIVs (not H9 or N2) could be amplified into one specific band, 667 bp for M gene. No specific bands of the same sizes were amplified from genomic materials of other avian pathogens. The detection limit of triplex PCR was 10^-2 ng/μL. The results of triplex PCR for detection of clinical samples were coincident with that of the viral isolation completely. Our results demonstrated that the optimized triplex PCR assay was quick, specific and sensitive for detection of AIVs especially H9 and N2 subtype AIV.

Key words: avian influenza virus; H9 subtype; N2 subtype; M gene; triplex PCR

CLC Number:

S852.65

XU Qian, XIE Zhi-xun, XIE Li-ji, LUO Si-si, HUANG Li, HUANG Jiao-ling, ZENG Ting-ting, XIE Zhi-qin, DENG Xian-wen. Development of a Triplex RT-PCR Assay for Detection of Avian Influenza Virus H9N2 Subtype[J]. , 2014, 41(11): 58-62.

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Development of a Triplex RT-PCR Assay for Detection of Avian Influenza Virus H9N2 Subtype

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