為建立一種快速準(zhǔn)確檢測H9亞型禽流感病毒基因的檢測方法，根據(jù)GenBank中H9亞型禽流感病毒血凝素編碼基因進(jìn)行熒光檢測引物和探針設(shè)計(jì)，建立了一種針對H9亞型禽流感病毒的熒光RT-PCR檢測方法。利用該方法進(jìn)行靈敏度和特異性檢測，并用本方法和國家標(biāo)準(zhǔn)中的熒光RT-PCR檢測方法，同時(shí)對臨床樣本進(jìn)行檢測。結(jié)果顯示，僅H9亞型禽流感病毒出現(xiàn)了正常熒光檢測曲線，而其他病毒及陰性對照均未出現(xiàn)；檢測靈敏度為RNA終濃度10-4 ng/μL（1.30×104 copies/μL）；臨床樣本檢測結(jié)果與國標(biāo)方法一致，符合率為100%。結(jié)果表明，該方法特異性強(qiáng)、靈敏度高，可用于H9亞型禽流感病毒檢測。
Establishment and Application of a Real-time RT-PCR Assay for Detection
of H9 Subtype Avian Influenza Virus
In order to establish a method for detecting H9 subtype avian influenza virus（AIV-H9）rapidly and accurately，the fluorescence detection primers and probes were designed and synthesized according to hemagglutinin encoding gene of AIV-H9 registered in GenBank，and a fluorescence PCR assay was thereby established，and its specificity and sensitivity were tested. Then by the established assay and the RT-PCR stipulated in national standard，some clinical samples were simultaneously tested. The results showed that the normal fluorescence detection curve only appeared in AIV-H9 rather than other non-specific viruses and negative sample；its detection limit was 10-4 ng/μL（1.30×104 copies/μL）of RNA；the detection result of clinical samples by established assay was consistent with the national standard，and the coincidence rate was 100%. In conclusion，the assay could be used to detect AIV-H9 due to its strong specificity and high sensitivity.
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