Fingerprinting method in a single microchip

Fingerprinting; Human DNA fingerprinting is currently almost exclusively performed based on short tandem repeat (STR) region identifcation. Because of the highly polymorphic nature of these repetitive sequences, very distinctive profles can be generated from minute amounts of DNA allowing discrimination of different individuals. Hence, a global consensus to use specific STR loci was agree upon, allowing straightforward comparison of profiles in databases around the world.

Te gold standard method for generating STR profiles; which was optimize about two decades ago and consists of a combination of multiplex polymerase chain reaction (PCR); also a subsequent fragment sizing using capillary electrophoresis (CE). Although delivering excellent results; so this method of STR profiling does have some drawbacks.

Fingerprinting analysis

This information be use to focus the investigation on certain suspects; also exclude innocent suspects at an early stage. An initial step would be to overcome some of the logistical steps of a traditional DNA fingerprinting analysis; also perform a rapid on-site DNA screening. Lab-on-a-chip (LoC) technology; which could address some of the limitations that hamper; so the conventional STR typing process and thereby provide fast, correct information.

This limits the number of repeats available for probe hybridization and increases the diference in Tm between unblock STRs of varying length. To detect the full range of common alleles; so up to three blocker sequences per locus are require. Therefore, a LoC capable of simultaneously amplifying; also melting more than reactions to be design and test. To allow a reference and crime scene sample to be analyze simultaneously a LoC capable of analyzing assays in parallel was design.

The miniature chip holds small, reaction cavities for parallel testing and an integrate aluminum heating system. Temperature monitoring was accomplish by a resistance temperature detector (RTD) deposit directly on the silicon surface. Te performance of this novel LoC to generate STR profles using HyBeacon technology was evaluate.

Detection of the alleles

All modify HyBeacon assays were test using conventional PCR amplifcation and melting analysis in a benchtop instrument before being transfer on-chip. Next, on-chip temperature uniformity during heating was assess over all cavities. Finally, complete STR proflies of three reference samples were generate on-chip and compare to of-chip results.

In this work the potential of an integrate silicon µPCR chip for the amplifcation of STR loci and detection of the alleles using a HyBeacon assays was demonstrate. A µPCR chip capable of performing 24 analyses in parallel is design and produce relying solely on standard semi-conductor fabrication processes.

Te potential of this µPCR chip to perform on-site DNA fnger printing was demonstrate by repeatedly and correctly producing STR profles of three reference samples. In addition to the previously develop D16S539, D18S51, D8S1179 and TH01 assays the generate STR profles also contain a sex identifer. A novel amelogenin gender determination assay was develop and optimize for on-chip analysis. Tis work is an important step towards the development of a handheld forensic analysis tool for on-site DNA fingerprinting