| Chinese Cherry is the perennial woody plant,belonging to genus Cerasus,subfamily Prunoideae,family Rosaceae.It has an early florescence,fresh and beautiful flowers,aromatic flavour,early fertile capability,with great value for food and ornament.In addition,Chinese cherry has wide adaptability and rich genetic diversity,which can be used as rootstock and breeding resources.So far,the researches mainly focused on phenotypic traits,cytology and molecular markers of the Chinese cherry germplasm resources.And cytology is the most controversial.In the previous study,only the karyotype analysis results of Chinese cherry were varied from different authors and the molecular cytology of Chinese cherries is still blank.In this study,the traditional karyotype analysis method was used to further explore the chromosome number and ploidy of Chinese cherry and the evolution trend.This study is mainly based on the karyotype analysis with the Pressed disc metnod and the Drop method,to discuss the karyotype evolution trend of Chinese cherry germplasm resources and the internal correlation;Explore chromosome in situ hybridization system by means of indirect and direct marker probe methods;Fluorescence in situ hybridization is established with 45 S rDNA and 5S rDNA as probes,to explore the distribution pattern of rDNA in Chinesee cherry's chromosome;Genome in situ hybridization system is established with self-genome and proximal species which are Cerasus avium,C.conradinae(2n = 2x = 16),C.serrulata var.Lannesiana(2n = 2x = 16)and C.campanulata as probes,to judgy the relationship between Chinese cherry and proximal species preliminarily;The homology of Chinese cherry chromosome group is preliminarily determined.These results were listed as follows:i Metaphase chromosome preparation Both pressed disc and drop methods are suitable for Chinese cherry chromosome preparation.By comparing their conditions,the press dics can be used for chromosome number and karyotype analysis,while drop method can be used for in situ hybridization for molecular cytogenetics.ii The ploidy level and karyotype of Chinese cherry There were three types of chromosome number and three ploidy levels in Chinese Cherry germplasm resources,which were tetraploid,pentaploid,and hexaploid with chromosome number of 32,40,and 48,respectively.Among them,cultivar from Yingjing is pentaploid,Fengxian germplasm is hexaploid.No diploids or triploids were found in this study.The karyotypes of Chinese Cherry are all "1A" type.The karyotype formula was mainly characterized by 2n = 4x = 32 = 32 m,and 2n = 4x = 32 = 28 m + 4sm for tetraploid;2n = 5x = 40 = 40 m for pentaploid;2n = 6x = 48 = 48 m for hexaploid.All these tested materials,Average arm ratio varied from 1.1 to 2.7,and length ratio varied from 1.28 to 2.17.The relative length of chromosomes of Chinese cherry type are mainly composed of M2 and M1,and less S chromosomes.iii Chromosomal in situ hybridization system The direct labeling system of the chromosomal in situ hybridization was selected on Chinese cherry.The probe concentration was about 1000 ng/?L,and the optimal hybridization period was from 15 to 18 hours.iv The chromosomal distribution of 45 S rDNA and 5S rDNA on Chinese cherry The chromosomes of Chinese cherry were analyzed by fluorescence in situ hybridization using rDNA as probe.There were detected eight 5S rDNA and six 45 S rDNA sites on Chinese cherry's chromosomes.There were 4 stronger singles of 5S rDNA on the short arm near centromere region and 4 weaker singles of 5S rDNA in the end of chromosomes shrot arms.There are 2 stronger singles at the end of chromosomes,2 weaker singles in the shrot arm near the centromere region and 2 weakest singles in the the secondary constrition.v The distribution of genomic DNA on Chinese cherry chromosomes We also conducted genomic in situ hybridization(GISH)analysis in Chinese cherry using the genomic DNA probes from Cerasus relative species.The Chinese cherry DNA probe generated a large number of signals dispersedly distributed through all chromosomes in Chinese cherry.The genomic probes of C.conradinae and C.campanulata(2n = 2x = 16)also gave strong signals on the terminal regions of 32 chromosomes,while C.serrulata var.Lannesiana(2n = 2x = 16)generated weaker signals located at the telomeric regions on each chromosome.The weakest hybridization signals from the C.avium mainly existed at the telomere region.Based on the hybridization signal number and intensity,GISH results suggested that the genome of C.conradinae and C.campanulata were most closely related to that of Chinese cherry,follwed by C.serrulata var.lannesiana,whereas C.avium was distanly related with Chinese cherry. |
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