The RT-PCR amplification of full length RNA of hepatitis a virus H2 strain.

A method for RT-PCR amplifying long fragments cDNA was established and the 7.4 kb full-length cDNA of Hepatitis A virus (HAV) H-2 strain was amplifyed by reverse transcription and polymerase chain reaction (RT-PCR). HAV H-2 was precipitated by anti-H-2 serum specifically, then the RNA of HAV H-2 was isolated from this precipitation by acid guanidinium hydrochloride-phenal-chloroform extraction. The first-strand of HAV H-2 cDNA was synthesised by reverse transcriptase without RNase H activity, then was amplified by PCR using the 32mer primer and the Tag DNA polymerase with Deep Vent DNA polymerase. For obtaining longer PCR products, it is necessary to prepare high quality RNA and employ the longer primers and special Tag DNA polymerase.