High-throughput S-SAP by fluorescent multiplex PCR and capillary electrophoresis in plants

The inherent replicative mode of transposition endows retrotransposons with considerable advantages as genetic tools in plant genome analysis. Here we present a high-throughput sequence-specific amplification polymorphism (S-SAP) method based on copia-like retrotransposons to fulfill the increasing desire of screening large numbers of samples in plants. Classic approach for digestion, ligation and pre-amplification was combined with optimized fluorescent multiplex PCR for simultaneously selective amplifying S-SAP fragments, and multiple S-SAPs were subsequently detected by capillary electrophoresis using ABI PRISM 3700 capillary instruments. Comparisons of results from multiplex PCR with simplex PCR, and from capillary electrophoresis with slab-gel electrophoresis demonstrated that this method is an efficient, economical, and accurate means for high-throughput and large-scale genotyping retrotransposon variation in plants. (C) 2004 Elsevier B.V. All rights reserved.