| Cymbidium grandiflorium, as the world famous "Orchid Star" in the Cymbidium orchid plants, was liked by most consumers with genetle flowery flavour and elegant belong to orchids and rich and colorful belong to tropical orchids. To provide genetic breeding and cultivation of scientific basis for Cymbidium grandiflorium, the botanical method and the principle and technique of SRAP molecular marker were used to research the morphological characteristics and genetic diversity of the germplasm resources of 42 Cymbidium grandiflorium species which come from South Korea, Japan, Australia, China Taiwan's and four wild species from China. The main results were indicated as follows:1. Cymbidium grandiflorium characteristics analysis showed that 46 tested varieties can be divided into two categories varieties. One was the wild species from China with the smaller and thinner flowers, and the more fresh and elegant flower color. The other category was Cymbidium grandiflorium species with the larger and denser flowers and more colorful. In the 42 tested varieties of Cymbidium grandiflorium species,4 Korean Cymbidium grandiflorium species were large flowers, while 17 Japanese Cymbidium grandiflorium species were mostly large flowers. As the different traits of different species, the hybrid between good traits has certain role in promoting the development of the flower industry, and be beneficial to the conservation and use of the Cymbidium grandiflorium genetic diversity.2. Cymbidium grandiflorium SRAP-PCR reaction system optimization showed that the best reaction system was:20μL volume, which included lxbuffer(10 mmol/LTris-HCl, pH 8.0,50 mmol/LKCl,2.3 mmol/L MgCl2,0.08% Nonidet P40), dNTP(every 200μmol/L), forward primer 33ng, reverse primer 33ng, DNA template 20ng, Taq enzyme 1.5U. Pre-amplification conditions was 94℃for denaturing 5min, responsed to the first 5 cycles at 94℃1min,35℃1min,72℃1min; followed by 30 cycles of annealing temperature up to 60℃, the final extension at 72℃5min,4℃saved. PCR was carried out in MG96G instrument of gradient PCR.3. SRAP analysis of Cymbidium grandiflorium germplasm resources showed that 34 pairs of SRAP primer combinations were screened out 29 pairs stable band-type and good polymorphism primer combinations. Among the 428 bands obtained 421 polymorphic bands, polymorphic ratio was 98.3%. The average number of DNA band per primer was 14.7. Germplasm similarity based on dice coeffcients among the cultivars ranged of 0.60 to 0.99, with an average of 0.76.29 SRAP markers based on amplification results, using NTSYS 2.10e software to calculate Dice coefficients of genetic similarity and the establishment of UPGMA dendrogram. The genetic similarity coefficient in 0.69 would be divided 46 for test material into four types:a species of orchid native species-prostrata Cymbidium faberi Rolfe; including three wild species from China, Cymbidium Kanran Makino, Cymbidium goeringii and Cybidium ensifolium. orchid native species; only hybrid orchid; includeed all 41 Cymbidium grandiflorium species. The result better revealed the genetic diversity and relationship in Cymbidium grandiflorium and primary species of Chinese orchid, and provided scientific basis for Cymbidium grandiflorium germplasm resource utilization and hybrid breeding parents choosed.4. Principal component analysis of Cymbidium grandiflorium germplasm resources showed that 46 tested materials were divided into five groups by origin:Group I included four primary species of Chinese orchid; GroupⅡincluded four general species from Korea; GroupIII included fourteen general species from Japan; GroupIV included twelve general species from Taiwan and Group V included fourteen general species from Australia. The result revealed that the genetic differentiation of species and hybrid species is both certain diference and inherent relevant with accompanied with increasing generations of offsprings, and better reflected the genetic diversity and relationship of Cymbidium grandiflorium germplasm. |
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