A PLANT DNA ISOLATION PROTOCOL SUITABLE FOR POLYMERASE CHAIN-REACTIONBASED MARKER-ASSISTED BREEDING

An important, but often limiting step in marker-assisted breeding is t he efficient isolation of plant DNA for polymerase chain reaction (PCR ) amplification. While there are many protocols for plant DNA isolatio n, 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 process ing in a 96-well microtiter plate. A single person fan isolate DNA and initiate PCR-based analyses for 96 individuals in one day. The DNA sa mples are suitable for several PCR-based procedures, including random amplified polymorphic DNA (RAPD), microsatellite (simple sequence repe ats), and amplified fragment length polymorphism (AFLP) analyses. Beca use the DNA is immobilized on a solid matrix and processed in 96-well plates, this protocol could be modified for robotic manipulation.