DNA Fingerprinting Of Prunus Mume Sieb. Et Zucc.

Prunus mume Sieb. et Zucc. is a famous ornamental tree species grown in China playing an important role in Chinese gardening and people's cultural life. The objectives of this study are to develop superior cultivars, and to effectively conserve and utilized the superior genetic resources, to develop methods of identification at molecular level for international registration and new variety registration of the species/varieties. 187 samples of Prunus mume and related species were analyzed by using AFLP markers with silver staining. DNA fingerprints were established for typical species/varieties, based on the AFLP techniques and the DNA fingerprints, the identification of Mei cultivars, the relationship between Prunus mume and related species and core collection of germplasm were studies with use of NTSYS 2.1 software. The main results were as follows:1. A Silver-staining AFLP system suitable for Prunus mume was developed, clearer and high resolution bands were obtained with appropriate handlings in sample collection, DNA extraction, restriction-ligase reaction, and double amplification as well as staining techniques throughout the experiment process. The techniques developed can be applied in normal molecular biology labs.2. With use of Mse I - EcoR I digest and primers, fingerprints of the 187 samples were obtained. With 7 combinations of primers, 373 loci, 83776 polymorphic bands were obtained with an average identifying percentage of 95.87%. The primer E-GA/M-CTC was taken as an example to conduct cluster analysis with use of NTSYS software to reveal the similarity among varieties. Results indicated that AFLP is highly effective for identification of Prunus mume varieties. However, varieties that are obviously different in morphology may have very similar fingerprints with certain primers, and those morphologically similar varieties may have large difference in fingerprints for some primers. The correlation between percentage of polymorphic bands and identification efficiency is low. Therefore, combination of AFLP techniques with morphological and enzyme techniques is essential for variety identification.3. Results of variety identification and phylogenetic relationship among plum varieties showed complex origin and evolution of Mei. Relatedness between some peaches and some Mei varieties were studied. The classification wassupported that Apricot Mei Branch was included in Mei.4. Four cultivars of ' Caishan Gongfen' , ' Qingming Wanfen' , 'DaGongfen' ,'Fenyu Gongfen' were proposed as the paternity of 'Meiren Mei' by the date of AFLP of 'Gong Fen' types, based on the AFLP fingerprints and the cluster analysis of Nei's genetic distance, SM, Dice coefficient.5. The core collection of germplasm of Prunus mume was established. 52 samples were selected out of 162 varieties as the core collection of germplasm based on the AFLP fingerprints and the cluster analysis of Nei's genetic distance. Genetic diversity of the pooled population was also tested. The selections mostly remained the genetic diversity and cultivar forms of total collection.Regarding international registration and new variety right protection of Prunus mume varieties, standardized fingerprints archive of Prunus mume varieties can be established. When varieties cannot be identified by morphological method, DNA fingerprints are therefore can be used for identification. However, the applicants for variety right should provide fresh somatic tissue of the plants.The constructed fingerprinting provided useful information not only for developing superior cultivars and new variety right protection but also for conserved and utilized the Mei genetic resources. The improved systems could be used for constructed standardized fingerprints archive of international registration of Mei cultivars, genetic map, marker-assisted selection and map based cloning in Prunus mume. The results, therefore, will produce important effects on genetic improvement of Prunus mume at the molecular levels.