Microspore Culture And Molecular Identification Of Self-incompatibility On Brassica Vegetables

Brassica vegetables are a kind of important commercial crops and the germplasm resources are extraordinary abundant in China. Most of them are cross-pollinated crops and have significant heterosis. Whereas it ususally takes about 5-8 years to develop a homozygous line using the traditional selection techniques, it is enough for 1-2 years using isolated microspore culture techeniques. At present, it is considered as a main method to produce commercial F1 hybrid cultivars using self-incompatible system in most Brassica vegetables, due to its wide presence. Nevertheless, it is extremely time-consuming for the identification of self-incompatibility strength and the types of S haplotypes. The objective here is to attempt to establish a rapid and highly efficient breeding system through combining the microspore culture and the molecular identification of self-incompatibility, further to improve the breeding procedure.Some factors during microspore culture and plant regeneration were explored for 3 kinds of Brassica vegetables (Chinese cabbage, purple flowering stalk and cabbage), a large number of embryoids and regenerated plants were obtained. Subsequently, we determined that the ploidy of plants derived from microspore culture, studied the genetic diversity, the strength and inheritance of self-incompatibility in DH populations. Moreover, we also investigated S haplotypes of several Brassica vegetables in China. The main results are as follows:1. Isolation microspore cultureAfter exploring 7 factors (genotype, developmental stage, activated charcoal, density of microspores, concentration of sucrose, heat shock and concentration of macro salts) influencing microspore embryogenesis, microspore embryogenesis was initiated in 73.3% of Chinese cabbage genotypes, 76.9% of purple flowering stalk and 50% cabbage. The treatments, such as the selection of 50-70% late uninucleate microspores, density of 1.5-2 buds/dish and addition of 0.3mg/ml activated charcoal, are helpful to microspore embryogenesis in Chinese cabbage and purple flowering stalk. Whereas the selection of 10-30% binucleate microspores, density of 1.5 buds/dish and addition of 0.1mg/ml activated charcoal are benefit to microspore embryogenesis in cabbage. The treatment of 13% sucrose is suitable for 3 kinds of Brassica vegetables. In addition, the induction frequency of embryoids significantly enhances in purple flowering stalk when combined heat shock pretreatment at 32℃ for 18h and a half of macro salts in the media.2. Plant regeneration from embryoidsAfter 4 factors (concentration of agar, embryoid age, low temperature and culture...