| There is a long history of rose breeding,and most modern Chinese roses are tetraploid varieties.It is very important to introduce the fine characters of Chinese ancient rose in the process of cultivating modern rose to promote the development of breeding.The Chinese ancient rose variety'Y18'(2n=2x=14)has the characteristics of continuous flowering,disease resistance and cold resistance,it is regarded as a representative model of the Chinese rose group.In order to successfully hybridize the'Y18' and modem rose,obtain new varieties with excellent germplasm,this study used colchicine to induce the stem of the 'Y18' sterile seedling,successfully obtained the tetraploid 'Y23',breaking the reproductive isolation between 'Y18' and the modern rose tetraploid variety 'H12'.The main research results are as follows:(1)The optimal disinfection method for diploid 'Y18' is to soak and disinfect with 2%NaClO for 10min,and it has the best proliferation effect in MS growth medium supplemented with 1.5 mg/L 6-BA and 0.05 mg/L NAA,the average proliferation coefficient is 3.48.The rootless seedling base was quickly inoculated in 0.5%IBA and inoculated in 1/4MS medium without any auxin to obtain the best rooting effect,the rooting speed was fast,the number of roots was large and relatively neat.The best induction effect was obtained by inducing the stem of the diploid 'Y18' sterile seedling obtained by tissue culture with 1%colchicine solution for 1h,and the induction rate was 26.1%.The survival rate of tetraploid was 53.3%after rapid propagation in tissue culture.(2)The ploidy of induced plants was identified by phenotypic traits observation,chromosome fragmentation,stomatal traits observation,chlorophyll content determination,POD activity,CAT activity,MDA content determination and cell flow cytometry identification.The results showed that compared with diploid plants,the plant type and flower diameter of induced plants increased significantly,and the leaf shape changed from lanceolate to oval.The results of chromosome fragmentation were 2n=4x=28,stomata significantly increased,chlorophyll a significantly increased,chlorophyll b and total chlorophyll content increased compared with diploid,but there was no significant difference.POD activity and MDA content were significantly different.(3)The 'H12' was used as the female parent,and the 'Y18' and the 'Y23' were used as the male parent for crossing.The results showed that 'H12'x'Y18' was not fruitful,and 'H12' x 'Y23' solidity rate was above 80%.The results of fluorescence observation showed that the pollen of 'Y18' and 'Y23'can germinate normally on the stigma of 'H12',but the pollen of `Y18' can not enter the style normally,and the pollen of 'Y23' can grow normally in the style of 'H12' and finally enter the ovary(4)After the seeds obtained from the 'H12' × 'Y23' were planted,50 F1 generations were harvested.Eighteen phenotypic traits of F1 generation were analyzed and eight principal components were extracted.Cluster analysis clustered 50 offspring into four clusters including ten groups(?-1,?-2,?-3,1-4,?-1,?-2,?-1,?-1,?-2 And ?-3).The first cluster(28 plants),the second cluster(3 plants),the third cluster(1 plant),the fourth cluster(18 plants),among them,the first three clusters of flowers are pink lines,which are more consistent with the color of the male parent,and the fourth cluster is red line,which is similar to the female parent.Correlation analysis between the traits showed that petal number was positively correlated with top leave length(.344)and aculeus amount(.352),and top leave width was positively correlated with top leave length(.466)and intemode(.441).(5)Four strains of 'YH03','YH14','YH37' and 'YH39' with good characters in F1 generation were selected for tissue culture and rapid propagation.The optimum medium for proliferation of'YH03','YH14' and 'YH37' was MS+1.5 mg/L 6-BA and 0.05 mg/L NAA,and the optimum medium for proliferation of 'YH39' was MS+1.5 mg/L 6-BA+0.10 mg/L NAA.Under the same rooting method,'YH37' had the highest rooting rate(86.67%).Among the four F1 generations,'YH37' had the best development,fast germination,high proliferation rate,good rooting effect and strong stress resistance.Therefore,In this experiment,'YH37' is the most suitable for tissue culture and rapid propagation,and the best culture conditions for other materials need to be further explored.This study used colchicine to induce diploid 'Y18' and successfully obtained its autotetraploid variety 'Yangjing Fen'.They were respectively hybridized with modern rose 'H12','H12' × 'Y18' was not fruitful,and 'H12' × 'Y23' has a high rate of solidity.The reason for the difference was analyzed by fluorescence observation of germination in vivo.Phenotypic traits of F1 hybrid plants of 'H12' and'Y23' were investigated and analyzed systematically.Four lines with better plant type were selected to establish tissue rapid propagation system. |
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