| Loop-mediated isothermal amplification (LAMP) method is a novel nucleic acidamplification method which amplifies DNA with high efficiency and rapidity underisothermal conditions with only simple incubators.A novel method of Avian reovirus detection by Loop-mediated Isothermal Amplification(LAMP) was reported using fluorescent reagent as the product judgment. With one set ofprimers containing six special primers targeting p10gene, the LAMP reaction could completewithin1hour at63℃. Sensitivity and specificity of this method was tested. The detect limitof the assay was102copies virus, which was100times more sensitive than traditional PCR.In the fluorescent reagent test, SYBR Green I and calcein reagent were respectively used inLAMP product judgement, and the result was consistent with agarose gel electrophoresis. Inthe detection of80clinical samples,68and55samples were respectively positive by LAMPand PCR assays.As the same time, According to the infectious bronchitis virus (IBV)5a+5b genesequences of6LAMP primer of8regions design a LAMP primer, develop a way onFluorescent Reagent in detection of avian IBV by LAMP. It can be completely responsed inisothermal (63℃) conditions by1hour. This method is of better sensitivity and specificity,can be detected1copy of positive restructuring plasmid,100times higher than normal PCR,and can’t detected other virus. Respectively with HNB reagent, its result is the same withagarose gel electrophoresis. In a clinical samples detection Of60, use LAMP and PCR testingpositive for IBV respectively of49and42.LAMP method is a rapid, accurate, and cost-effective diagnostic method for infectiousdiseases. Furthermore, the resμLt can directly obtain with the use of calcein dyes. Given themany advantages of LAMP, the paper established ARV-LAMP and IBV-LAMP method aresuitable for promotion.The method are expected to detect Avian reovirus and infectiousbronchitis virus in field situations. |
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