Bioassay for specific DNA sequences using a non-radioactive probe.
A novel method for detecting specific DNA sequences is described. The method uses a non-radioactive DNA probe, called a probe-vector, that can transform competent Escherichia coli cells at high efficiency only when it has hybridized to a specific DNA target, thus forming a circular, double-stranded, plasmid-like molecule. The probe-vector carries a plasmid origin of replication and a gene that confers antibiotic resistance on transformed E. coli. The output of the assay--colored bacterial colonies on an agar plate--is quantitative and proportional over a wide range of target concentrations. The utility of the probe-vector method for detecting hepatitis B virus (HBV) DNA in human serum is demonstrated. The assay can detect as little as 0.1 pg HBV DNA. The presence of an internal standard monitors DNA recovery and E. coli transformation efficiency for each sample. The assay has the potential to simultaneously measure the DNA of two or more pathogens within the same clinical sample.
D001681:Biological Assay; D004247:DNA / Q000032:analysis*; D004279:DNA, Viral / Q000032:analysis; D005822:Genetic Vectors; D006509:Hepatitis B / Q000175:diagnosis; D006515:Hepatitis B virus / Q000235:genetics; D006801:Humans; D010957:Plasmids; D014170:Transformation, Genetic
D004279:DNA, Viral; D004247:DNA