A Plant DNA Isolation Protocol Suitable for Polymerase Chain Reaction Based Marker‐Assisted Breeding

Abstract

An important, but often limiting step in marker‐assisted breeding is the efficient isolation of plant DNA for polymerase chain reaction (PCR) amplification. While there are many protocols for plant DNA isolation, they tend to be time consuming or difficult to use on many samples simultaneously. We have optimized an alternative approach to plant DNA isolation that immobilizes DNA on a solid matrix, followed by processing in a 96‐well microtiter plate. A single person can isolate DNA and initiate PCR‐based analyses for 96 individuals in one day. The DNA samples are suitable for several PCR‐based procedures, including random amplified polymorphic DNA (RAPD), microsatellite (simple sequence repeats), and amplified fragment length polymorphism (AFLP) analyses. Because the DNA is immobilized on a solid matrix and processed in 96‐well plates, this protocol could be modified for robotic manipulation.