title: Automated extraction and amplification of DNA from whole blood using a robotic workstation and an integrated thermocycler.
abstract: Growing knowledge of the genetic basis of inheritable diseases has resulted in a rapidly increasing demand for DNA mutation analysis. Current methods are reliable and suitable for low-throughput mutation analyses, but are unable to cope with the increasing demand for genetic analyses, necessitating the development of new, fully automated and reliable methods. We developed a semi-automated method for DNA mutation analysis by integrating a thermocycler into a robotic pipetting workstation. DNA was extracted from 84 samples of 10 microl of EDTA-treated whole blood using magnetic beads within 2 h. Directly after isolation, the DNA was automatically transferred to an integrated thermocycler for amplification. Our semi-automated method proved to be reliable and robust, showing unambiguously interpretable PCR signals without occurrence of contamination. It is also faster than conventional manual methods. Only a brief manual intervention is required to remove and refit the seal of the PCR plate. This semi-automated assay is a step forward in the development of fully automated assays for DNA mutation analysis.
authors.affiliation: 424 Department of Paediatrics, University Hospital Nijmegen (St Radboud), P.O. Box 9101, 6500 HB, Nijmegen, The Netherlands.
authors.forename: M L
authors.initials: ML
authors.lastname: Smit
authors.forename: B A
authors.initials: BA
authors.lastname: Giesendorf
authors.forename: S G
authors.initials: SG
authors.lastname: Heil
authors.forename: J A
authors.initials: JA
authors.lastname: Vet
authors.forename: F J
authors.initials: FJ
authors.lastname: Trijbels
authors.forename: H J
authors.initials: HJ
authors.lastname: Blom
mesh_terms: D004247:DNA / Q000097:blood* / Q000302:isolation & purification*; D006801:Humans; D042965:Methylenetetrahydrofolate Reductase (NADPH2); D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000097:blood / Q000235:genetics*; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000295:instrumentation*; D012371:Robotics
chemical_list: D004247:DNA; D000587:Oxidoreductases Acting on CH-NH Group Donors; D042965:Methylenetetrahydrofolate Reductase (NADPH2)