Sequence variation of chloroplast DNA that involves EcoRI and ClaI restriction site polymorphisms in soybean

Restriction fragment length polymorphisms (RFLPs) of EcoRI- and ClaI-digested chloroplast DNA (cpDNA) within the genus Glycine subgenus Soja were characterized. Two mutations were found to be responsible for the EcoRI and ClaI restriction site polymorphisms, and both were located in a region in which many ribosomal protein genes are clustered. This region is within the large single copy region of cpDNA and is located close to an inverted repeat. The locations of restriction sites of EcoRI and ClaI in the cpDNA region were analyzed by DNA gel-blot analyses and PCR amplification, which were followed by sequencing analyses. The EcoRI site polymorphism was found to have occurred in the intergenic spacer between rps11 and rpl36, while the ClaI site polymorphism was located within the 3' part of the coding region of rps3. The mutations that cause EcoRI and ClaI polymorphisms were both found to be single base substitutions. In addition to these polymorphisms, novel sequence variations in soybean cpDNA were detected near the sites of these mutations. Previously, it was shown that cultivated soybeans could be classified into three groups (I, II, and III) based on their cpDNA RFLPs. A comparison of the cpDNA sequences of soybeans in the present study was consistent with the notion that the cpDNA of group II soybeans is an intermediate between the cpDNAs of groups I and III.