Studies On The Biological Characters And New Germplasm Creation Of Some Staminate Germplasm In Diospyros L.native To China

Japanese persimmon(Diospyros kaki Thunb.) belongs to the Diospyros genus, Ebenaceae Family.There are abundant Japanese persimmon genetic resources in China and many of them are valuable.Some new staminate Diospyros spp. germplasms have been recently collected in Dabieshan Mountains,though the study on the basic biological characters and the breeding value of them was not much made abroad.Upon some staminate Diospyros spp.germplasms,our study focused on five aspects:the first was to investigate the growing stages,morogology of leaves and floewers,pollen germinability in vitro,pollen-stigma compatibility,and the ploidy. The second was valuated naturally occurrence rate of giant pollen and the cytological mechanism responsible for 2n pollen formation of these staminate germplasms by observing the pollen mother cells meiosis.The third was to establish the system of of zygotic ovules culture in vitro and find the optimum stage for embryo rescue in Japanese persimmon.The fourth was intra-specific cross among Mopanshi and Luotiantianshi with staminate Diospyros spp.germplasms,and other two pollination-constant and non-astringent(PCNA) type of Chinese with Luotiantianshi. The fifth was to study in vitro chromosome doubling during embryo culture and shoot apices culture of Luotiantianshi with two anti-microtubule herbicides and colchicines. The major results are indicated as follows:1.Flow cytometry analysis showed that as most oriental persimmon(D.kaki Thunb.),Male type No.1,Male type No.2,Male type No.3,Male type No.8,Male type No.9 and Male type No.10 were hexaploid.2.The phenology of these genotypes was similar with other varieties of Diospyros kaki in Wuhan,and the exterior morphological characteristic of leaves and floewers was distinct among these plants.And they had some valuable pollen characteristic,such as great pollen number,high pollen germinability in vitro,good pollen-stigma compatibility between six Diospyros spp.staminate genotypes and two female parents,'Maekawa Jiro' and 'Mopanshi'. 3.The pollen mother cells(PMCs) meiosis of staminate Diospyros spp. germplasms finished at the end of March in Wuhan.It took short time to finish what happened from metaphaseⅠto telophaseⅡ,and the synchronization of meiosis were greatly different in different genotypes,floral branches from the same genotype,and even anthers of the same flower.The first division during the meiosis of PMCs was found to proceed normally.Several abnormal types of spindles,including fused, tripolar spindles and abnormity-disintegration of sister chromatid were detected at metaphaseⅡ,which formed the dyads and triads,produce 2n pollen in the end.The frequencies of giant pollen,dyads and triads detected in all accessions were 0-2.3%, 0.8%-2.2%and 2.3%-4.3%,respectively.The correlation(r=0.98**) between the frequency of giant pollen and the frequency of dyads and triads was highly significant, which strongly suggested that giant pollen was 2n pollen.Most 2n pollen produced by tripolar spindles and fused spindles was equivalent to first division restitution(FDR) gametes,while the minority 2n pollen produced by abnormity-disintegration of sister chromatid was equivalent to second division restitution(SDR) gametes.4.The optimum stage for immature ovules culture was 25 days after pollination (DAP).MS(1/2N) media containing 1.00m g·L^-1IAA and 2.20 m g·L^-1ZT was the optimum medium for inducing ovules germination.At 40 DAP,embryos were at globular stage.At 50 DAP,heart and torpedo embryos were found.Embryos were at the stage of cotyledon from 60 DAP.The rate of germination of globular embryos at 40 DAP(29.3%) was significantly less than embryos at other stages,and all embryos after heart stage germinated at 100%.Results of the plantlets regeneration percentage, the height of seedlings and number of seedling roots at 60 DAP was significantly better than other four stages.So the optimum stage for embryo rescue in persimmon was from heart stage to cotyledon stage which was 50～60 DAP in Wuhan,Hubei province.5.137embryo rescue seedlings were obtained from 6 crosses when Mopanshi as female parent,staminate Diospyros spp.germplasms as male parent,including:7 seedlings from Mopanshi×Male type No.1,28 seedlings from Mopanshi×Male type No.2,72 seedlings from Mopanshi×Male type No.3,5 seedlings from Mopanshi×Male type No.8,2 seedlings from Mopanshi×Male type No.9,23 seedlings from Mopanshi×Male type No.10.Some of seedlines were established in the vinevard,and hybrids generally grew abnormally.The morphology of the hybrids inclined predominantly to Mopanshi morphology character.Moreover,228 embryo rescue seedlings were obtained from 3 crosses when Mopanshi,Eshi 1,Baogaitianshi as female parent,Luotiantianshi as male parent,including:185 seedlings from Eshi 1×Luotiantianshi,43 seedlings from Baogaitianshi×Luotiantianshi,no seedlings from Mopanshi×Luotiantianshi.108 seeds were obtained from 3 crosses when staminate Diospyros spp.germplasms as male parent,including 14 seeds from Luotiantianshi×Male type No.2,29seeds from Luotiantianshi×Male type No.3,64seeds from Luotiantianshi×Male type No.10.6.Antimicrotuble could induce polyploidy by treating 'Luotian Tiashi' persimmon in vitro shoot-tips and embryos.The results indicated that oryzalin was the most effective inducers,the optimal treatments was 200μmol/L oryzalin for 2or 4 d.No polyploidy was induced by colchicine treatments.And the most number of dodecaploid plantlets was obtained by treatment with 100μmol/ L trifluralin for 30days,and the percentage of dodecaploid reached 19.4%.Stomata of the mutated plantlets were significantly larger than those of the normal plantlets,but the density of stomata was reduced significantly.And the leaf weight of the mutated plantlets were higher than that of normal plantlets The nuclear DNA content of the mutated plantlets was twice as much as normal hexaploid plantlets.