Creation And Identification Of Ploidy Materials In Cucumber (Cucumis Sativus L.)

The narrow genetic basis and limited genetic diversity of germplasm resources are the main bottlenecks of genetic and breeding researches of cucumber (Cucumis sativus L., 2n=14), more and more emphases have been focused on the ploidy breeding. Molecular and cytogenetical studies could be promoted by selecting different ploidy germplasm resources (euploid and aneuploid) in cucumber. Furthermore, haploid culture technique could accelerate process of breeding through using homozy (haploid or doubled haploid) coming from haploid culture. Thus, the experiment is very important for theory research and breeding application.In our study, on the one hand, the germinating seeds of cucumber were treated with colchicine solution to induce the chromosome doubling. On the other hand, through unpollinated ovary culture and isolated microspore culture, we observed the explants of different genotypic materials’development process, and studied main factors affecting the haploid culture.1. Induction and identification of different ploidy materials in cucumberThe germinated seeds were treated with 0.4% colchicine solution, eight autotetraploidy plants (2n=28) and three aneuploidy plants (2n=16,19, or 27) were obtained in our study. Autotriploid plants (2n=21) were produced from the cross of autotetraploid and diploid. FISH result further indicated the ploidy level. There were different characteristics of morphology and leaf stoma in different ploidy plants. The morphologic characteristics and leaf stoma under electron microscope scanning could be used as an aid for identifying the ploidy of cucumber. This study identified chromosomal constitution of the aneuploidy plants with the chromosome number of 16. FISH result confirmed that this aneuploidy plant was assigned as tetrasome polyploidy with two extra chromosome 1 of cucumber (2n=14+2). These results confirmed that colchicine could directly not only induce autopolyploids, but also induce a variety of aneuploids.2. Optimization of unpollinated ovary culture system in cucumberTwo genotypes of cucumber ’P02xL8’,’P02xBaiyu’were used to study the effects of several factors to unpollinated ovary culture in cucumber, these factors contained sampling time, pretreatment time and additive. The results showed that the best sampling time of materials was 0-2 d before flowering. The high temperature pretreatment and additive were necessary to the in vitro culture of unpollinated ovary in cucumber, the sliced ovaries were vaccinated on medium MS+0.04 mg/L TDZ+20 mg/L AgNO3 in the laboratory, then they were incubated under 35℃ for 3 d in the dark to induce in vitro gynogenesis.3. Optimization of isolated microspore culture system in cucumberThirty genotypes of cucumber were used to study microspore developmental stages and the correlation with morphological characteristics of flower buds. Meanwhile, we studied the effects of sterilization method, genotype and hormone combination on isolated microspore culture in cucumber. The results showed that there were relationships between the microspore periods and the morphological characteristics that were the length or diameter of flower buds and the color of anthers in cucumber. The length or diameter of flower buds could be used as a basis to take sample. Cucumber was characterized by an asynchrony of pollen development within a signal flower bud or anther. The flower buds 0.6-0.8 cm in length and the light green of anthers were determined to contain microspores at the late-uninucleate stage mostly. Different treatments of genotype and hormone combination had the same observations; the results indicated the best sterilization method was 5% NaClO315 min; the studies had showed that part of isolated microspores swelled after incubating 3 d. Callus or embryoid were not observed in NLN liquid medium after culturing a period of time.