Classification of rice germplasm. I. Analysis using ALP and PCR-based RFLP

The potential of using a PCR-based approach to detect DNA polymorphism for rice germplasm classification was compared with that of Southern-based RFLP analysis. Thirty-five Iranian rice varieties were studied along with 2 typical Indica and 3 typical Japonica varieties. Thirteen mapped RFLP markers were used as hybridization probes against Southern blots containing digests of one restriction endonuclease; 12 of the 13 probes detected polymorphism in the varieties. Fifteen sets of oligonucleotides derived from sequences near the ends of the same probes and of two other mapped probes were used as primers for PCR amplification of total genomic DNA of the varieties. Amplicon length polymorphisms (ALPs) were detected with 6 of the 15 sets of primers. To identify additional polymorphism, the PCR products were digested with nine different restriction endonucleases recognizing 4- or 5-bp DNA sequences and analyzed by gel electrophoresis in agarose and polyacrylamide. RFLPs were detected for 11 sets of primers, due to point mutations and to addition/deletion events that were too small to be detected as ALPs. Because PCR products are easily generated and may be analyzed in detail through the use of restriction endonucleases that cut rice DNA frequently, PCR-based RFLP analysis is a useful tool for the classification of rice germplasm.