Studies Of Classification On Germplasm Of The Male Plants Of Ginkgo Biloba L.

Ginkgo biloba L., a dioecious plant,is a native of China with a rich resources in which the genetic constitution and diversity require understanding promptly and the classification need researching for its male plant germplasm resources because of its special importance at the afforest in garden, the allocation of pollenation trees, the cultivation for leaf-purpose, the development and utilization of pollen. In this study, the cultural history, the condition of growth and development, the characters of botany and economy, and the phenological period of 63 individuals of male plant distributing in main production districts of Yangzhou, Taixing and Jiangyan etc. in Jiangsu province were investigated, observed and measured, respectively. At the same time, young leaves were sampled from the top of shoots at 30 days after budding in the male plants for two years running for extracting genome of DNA in leaf with slightly improved CTAB, PCR amplification and primer selection,then, the results of ISSR marker, the measurements of the average area of single leaf, SLFW,SLDW of each male plant were used to take a cluster analysis with MATLAB based on Nei's distance and Ou's distance, respectively. The results indicated as follows:1. Out of 100 ISSR primers, 17 primers amplifying more polymorphic sites were selected, the sequences of which were(GA)₈C,(GA)₈A,(AC)₈G,(CT)₈GC,(CT)₈GG,(CT)₈RG,(AC)₈SG,(AC)₈GA,(AC)₈GG,(AC)₈YA,(AC)₈YG,(AG)₈ ST,(TC)₈RA,(TC)₈YT,(TC)₈RT,(CA)₈GT,(CA)₈AG., respectively;2. 115 clear bands from 17 primers, average 3¹0 ones from each primer after amplification, the fragments of DNA sequence with 100²400 bp, in which there were 59 polymorphic sites in total, were got. The percentage of polymorphic bands (PPB) was 48.57%, in which the PPB of (AC)8G,(AC)8SG,(AC)8GG were even more than 75%, and the PPB of the individuals of male plants were 32.79%⁴8. 10%;3. Within 63 individuals of male plant in this experiment, the percentage of hereditary diversity (Hsp%)of total samples was 18.8461%,and the Nei's distance of them were 0.0520⁰.3245. The genetic similarity coefficients were 0.6755⁰.9480. Based on complete linkage, the dendrogram of amplified results was constructed, in which when the threshold value of 0.2739 was taken, 63 male plants could be divided into 2 major groups including 23 and 40 individuals respectively,when 0.245 was taken as a threshold value, the above 2 principal types could be divided into 5 sub-groups, the former consisted of 2 sub-groups including 10 and 13 individuals, and the latter consisted of 3 sub-groups including 8,14 and 18 individuals respectively,which showed that the male plant of Ginkgo had a rich hereditary diversity, complex hereditary structure and background;4,In this experiment,the samples of male plant from the different areas were uniformly distributed in the two major groups and five sub-groups, which showed that even though the same type of male plant distributed in different areas, those male plants performed as a near genetic relationship. The genetic distance within 59 male plants from Yangzhou and Taixing showed shorter, which could be speculated that some of male plants were propagated by grafting scions exchanged each other between both districts. 4 male plants from Jiangyan, with a near genetic relationship, distributed together in the second group, which gave a hint that those male plants maybe come from the same germplasm. 28 male plants with age of 20～700 years in Yangzhou, showed a larger differences in genetic distance, in which there was a shorter genetic distance and a near genetic relationship among the male plants with age of over 200 years and the ones planted near to them, which may be explained that the old male plants were propagated by seedlings with different genes in different years, and the younger male plants developed in morden times were grafted by the scions from local male trees;5. 28 male plants in Yangzhou, after taking a further cluster analysis based on Nei's distance, the Nei's distance of which was 0.0520⁰.275,could be divided into 2 major groups including 16 and 12 individuals respectively when the threshold value of 0.25 was taken. When 0.245 was taken as a threshold value, the above 2 principal types could be further divided into 5 sub-groups, the former consisted of 3 sub-groups including 4, 6 and 6 individuals, and the latter consisted of 2 sub-groups including 6 and 6 individuals respectively;6. The average area of single leaf, SLFW,SLDW of 28 male plants in Yangzhou, the variation coefficients of which were 17.71%,15.99% and 17.62% respectively, were used to take a cluster analysis based on Ou's distance. Those male plants, the Ou's distance of which was 0.08²4.5,could be divided into 2 major groups including 15 and 13 individuals respectively when the threshold value of 24 was taken. When 16 was taken as a threshold value, the above 2 principal types could be further divided into 4 sub-groups, the former consisted of 2 sub-groups including 2 and 13 individuals, and the latter consisted of 2 sub-groups including 2 and 11 individuals respectively;7. The results indicated that after comparison of ISSR marker with cluster analysis of average area of single leaf, SLFW and SLDW, the allocation of 28 male plants in Yangzhou according to the relative indices of vegetative growth wasn't completely respondent to ISSR marker, which showed that the characters of botany and economy for male plants of Ginkgo biloba L. were both the results of gene expression and the results of comprehensive effects from environmental conditions, reproductive ways, plant age, nutrient condition, development stage etc.;8,ISSR marker, the rapid, simple, reliable, more polymorphic characteristics of which have been fully applied in this study, is suitable to the phylogenetic classification on the male plant germplasm resources of Ginkgo biloba L.. The results of both ISSR marker and cluster analysis provide a theoretical basis and technological support for enlarging sampling range and for combination of qualitative characters and quantitative characters needed by development and utilization of the male plants of Ginkgo biloba L. to make a further classification and denomination.