No abstract

We describe a unique human DNA resource forming part of the Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC), a longitudinal cohort study involving 14 000 children and their families living in a geographically defined area of England. The DNA bank will underpin the search for associations between genetic polymorphisms and common health outcomes. The opportunities to collect blood samples suitable for DNA extraction are necessarily limited, and the samples themselves have often been treated in different ways and have varied storage histories. With the need to maximise yields, the choice of DNA extraction method is critical to the success of the bank and we have investigated the suitability of various commercial and in-house methods of DNA extraction. Various steps have been taken to minimise errors in sample address and identification, including the use of a pipetting robot for dilution and transfer of samples between 96-well arrays to provide aliquots suitable for PCR. The robot has been programmed to cope with concentrated viscous DNA solutions.

No abstract

Allelic variation at the galactose-1-phosphate uridyltransferase gene undoubtedly plays a role in defining the biochemical and clinical phenotype. However, clinical galactosaemia is a complex trait in which multiple developmental and metabolic pathways are involved. Ultimately the phenotype is beyond the control of the single gene itself.

We describe a rapid and simple method for phenylketonuria genotyping which identifies five point mutations within exon 12 of the human phenylalanine hydroxylase gene. The method involves PCR amplification of the target exon and hybridization with a PCR-amplifiable synthetic DNA (universal heteroduplex generator, UHG). The UHG contains identifiers consisting of nucleotide substitutions and/or deletions, contiguous with known mutation sites within the target exon. DNA heteroduplexes are resolved by nondenaturing polyacrylamide minigel electrophoresis. Individual mutant genotypes are identified by characteristic banding patterns, in either homozygous or heterozygous states. The method may potentially be applied to rapid genotyping of any mutation or series of mutations within PCR-amplifiable genetic material.