Rapid DNA fingerprinting to control for specimen errors in HIV testing by the polymerase chain reaction

Abstract

Variable-number-tandem-repeats (VNTRs) are highly polymorphic and provide informative genetic markers for distinguishing between individuals. We have used PCR amplification of VNTR locus pMCT118 to identify mislabelled specimens submitted for HIV PCR testing. The method is rapid, can be applied to large numbers of samples and eliminates the need for radioactive probes. DNA samples (10 ng) are amplified for 25 cycles using fluorescence-labelled oligonucleotide primers (blue dye). An aliquot of the PCR product is then combined with an internal lane size standard (labelled with a red dye), electrophoresed through a 2% agarose gel on an automated fluorescence DNA fragment analyser and the size and quantity of the fragments determined automatically relative to the internal standard. Fifteen alleles, ranging in size from 398 tp 709 bp were readily identified in a random sampling of DNA from 63 unrelated HIV-infected patients. Fragment size was reproducible and corresponded to alleles containing from 16 to 35 repeats of a 16 bp unit. VNTR genotyping will prove useful for resolving discordant results due to specimen mix-up and ensuring that the correct samples have been analyzed.