Development of multiplex PCR markers to distinguish evergreen and deciduous azaleas

摘要

We successfully developed new PCR-based markers for analysing the DNA inheritance patterns of vigorous inter-subgeneric hybrids of azalea. The hybrids were between evergreen azaleas (Rhododendron nakaharae and its hybrids) and fragrant deciduous azaleas (Rhododendron arborescens and Rhododendron viscosum). We determined the sequence of the nuclear internal transcribed spacer (ITS) region in evergreen and deciduous azaleas, and found a deletion of five base pairs (bp) in evergreen azaleas compared with deciduous azaleas. Sequences for the chloroplast matK-trnK region were obtained from GenBank. Primers for the multiplex PCRs were designed using these sequence data. In the multiplex PCR of the ITS region, evergreen azaleas showed a specific band of 140 bp and deciduous azaleas showed a specific band of 800 bp. In the multiplex PCR of the matK-trnK region, evergreen azaleas showed a specific band of 560 bp and deciduous azaleas showed a specific band of 300 bp. These results indicate that multiplex PCR methods are useful to distinguish evergreen and deciduous azaleas easily and rapidly.