Polymerase chain reaction-free variable-number tandem repeat typing using gold nanoparticle-DNA monoconjugates.

In this work, we report a novel polymerase chain reaction (PCR)-free variable-number tandem repeat (VNTR) typing method using a T-shaped gold nanoparticle-DNA monoconjugate, called the "watching-gene assay". The T-shaped DNA probe was synthesized by "click" chemistry and linked with the gold nanoparticle to form the gold nanoparticle-DNA monoconjugate (a VNTR probe). Through a simple annealing and ligation reaction of the VNTR probe on a synthetic DNA template mimicking the human D1S80 VNTR locus, the number of tandem repeat units could be deciphered by counting the self-assembled gold nanoparticles. The number of tandem repeat units could be identified with more than 50% yield if the repeat number was less than four. In the case of the real human genomic DNA, the 18 repeat unit number could be successfully revealed by observing the 18-gold-nanoparticle cluster, which exactly corresponded to the number of tandem repeats of the real sample. Our "watching-gene assay" is rapid, simple, and direct for data interpretation, thereby providing an advanced PCR-free genetic polymorphism analysis platform.