New triphosphate conjugates bearing reporter groups: labeling of DNA fragments for microarray analysis

A simple and convenient method for incorporation of fluorescent or ligand groups into 3'-termini of DNA fragments is proposed. A set of triphosphoric acid monoesters bearing fluorescent groups or biotin attached to the triphosphate fragment through linkers of different lengths and structures was synthesized. All the compounds were substrates for calf thymus terminal deoxynucleotidyltransferase and were used for incorporation of marker groups into 3'-termini of DNA fragments. The compounds were successfully applied for DNA labeling during post-PCR target preparation for microarray analysis.     

HLA-DQA1, AB0, and AMEL genotyping of biological material with biochips

A method for genotyping of biological material for AB0, HLA-DQA1, and AMEL loci is described. The method utilizes allele-specific SNP typing with th e help of hydrogel biochip technology. Amplified fluorescentlylabeled fragments of genes were hybridized with SNP-specific DNA probes immobilized on a biochip. The alleles of the gene in a sample were determined according to the distribution of the fluorescent signal. The minimal amount of biological material required for the assay is 100 pg of DNA. The biochip assay was used to analyze 442 DNA specimens of the Eastern Slavic population of Russia and to determine the allelic frequencies of AB0 and HLA-DQA1 loci. The feasibility of genotyping the biological traces of investigative value, such as cigarette butts, sweat traces on paper, and swabs of the lip of the glass, was demonstrated. This assay is applicable in forensic studies. The significance of identification for the used loci is 99.6%.