Automated extraction and amplification of DNA from whole blood using a robotic workstation and an integrated thermocycler.
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.
Smit
M L
ML
424 Department of Paediatrics, University Hospital Nijmegen (St Radboud), P.O. Box 9101, 6500 HB, Nijmegen, The Netherlands.
Giesendorf
B A
BA
Heil
S G
SG
Vet
J A
JA
Trijbels
F J
FJ
Blom
H J
HJ
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
D004247:DNA; D000587:Oxidoreductases Acting on CH-NH Group Donors; D042965:Methylenetetrahydrofolate Reductase (NADPH2)