A method for detection of hepatitis B virus pre C G1896A gene mutant by PCR amplification blocking associated with fluorescent probe / 临床检验杂志

ABSTRACT

Objective To investigate and evaluate the method of PCR amplification blocking associated with fluorescent probe for the detection of pre-C region of HBV G1896A gene mutation.Methods The primers were designed based on the mutation of HBV DNA 1896 locus.The 3′end of primers was at 1896 site,and it was complemented with the base sequence of mutation template of 1896 site.The mismatching bases were separately introduced into the second and the third base of the primer by inverse counting from the 3′end for increasing the specificity of reaction.Results The PCR amplification for wild plasmid with the mutant primer showed an effectively blocking,but not showed blocking for the mutant plasmid (G1896A).The sensitivity of detection for the mutant plasmid was 5?103 copies/ml.Ninety-five cases of HBV-positive serum was selected randomly and amplified with the mutant primer,and 8 cases were positive HBV G1896A gene mutant(mutant rate of 8.4%).Conclusion The amplification blocking associated with fluorescent probe for the detection of HBV G1896A gene mutation is a effective,convenient method for the detection of clinical samples.