Dual RT-PCR (reverse transcription-polymerase chain reaction) method for quickly identifying duck hepatitis A virus serotype

Abstract

The invention relates to a dual RT-PCR (reverse transcription-polymerase chain reaction) method for quickly identifying duck hepatitis A virus serotype, which comprises the following steps: respectively synthesizing specific primers SEQ1 and SEQ2 for duck hepatitis A virus type 1 and specific primers SEQ3 and SEQ4 for duck hepatitis A virus type 3; by using RNA (ribonucleic acid) of the sample to be detected as a template and SEQ1 and SEQ3 as the primers, carrying out RT to obtain the virus cDNA (complementary deoxyribonucleic acid); and by using the two pairs of primers as dual primers and virus cDNA as a template, carrying out PCR reaction in the same reaction system, thereby simultaneously completing the quick identification of the duck hepatitis A virus type 1 and duck hepatitis A virus type 3. On the basis of the principle of the RT-PCR technology, the invention establishes a dual RT-PCR detection method according to the correspondence between the duck hepatitis A virus genotype and serotype as well as the characteristic that China mainland only has two serotypes of duck hepatitis A virus. The method is simple and quick, has the advantages of high specificity and high sensitivity, and can be used for quick serotype identification of duck hepatitis A virus type 1 and duck hepatitis A virus type 3.

Description

(1) Technical field [0001] The invention relates to a double RT-PCR method for quickly identifying duck hepatitis A virus serotypes, belonging to the field of virus molecular biology. (2) Background technology [0002] Duck hepatitis A virus (DHAV) includes three genotypes and serotypes: duck hepatitis A virus type 1 (DHAV-1), duck hepatitis A virus type 2 (DHAV-2) and duck hepatitis A virus type 3 (DHAV-3). ), almost no cross-protection among the three serotypes. In recent years, the pathogens of duck viral hepatitis in my country have become more and more complicated. Studies have shown that DHAV-2 has not been detected in mainland China, but mixed infections of DHAV-1 and DHAV-3 are common, causing serious economic losses to the duck industry. [0003] At present, the identification of duck viral hepatitis mainly depends on epidemiological investigation, clinical symptoms and pathological changes, but duck viral hepatitis caused by different duck hepatitis A viruses is v...

AI Technical Summary

Problems solved by technology

At present, the single and mixed infections of DHAV-1 and DHAV-3 are the most common in duck populations in my country. Methods for detecting duck hepatitis A virus using multiple RT-PCR methods have been established, but these methods have low sensitivity (Huang Qiuxue et al., Establishment of dual RT-PCR detection method for duck hepatitis A virus genotype A and C, 2012, 34(2): 120-123, the lowest detection sensitivity: DHAV-A is 4.98×10 4 copies / μl and DHAV-C is 1.68×104 copies / μl; He Ranya et al., Establishment of RT-PCR detection method for differential identification of type Ⅰ duck viral hepatitis virus and new duck hepatitis virus, Heilongjiang Animal Husbandry and Veterinary Medicine, 2009, 3:14 -16, the lowest detection sensitivity: DHAV-A is 0.8ng / μl and DHAV-C is 1.0ng / μl), or the shortcomings that cannot be applied to the detection of all prevalent strains in current production (Kim et al., 2008, Differential diagnosis between type-specific duck hepatitis virus type1 (DHV-1) and recent Korean DHV-1-like isolates using a multiplex polymerase chain reaction, 2008, Avian Pathology, 37:171-177)

Images