| Anthers from thirty-four genotypes of potato were cultured in the experiment. The effects of genotype, types of hormone on callus formation and differentiation in anther culture were analyzed; Factors that influence callus induction, such as developmental stage of pollen, pretreatment, sucrose concentration during the culture process, were investigated using seven genotypes of Ne16, Kexin13, VA, Neishu7, Zhongshu5, Sebago and Luyin 1. Factors that influence callus induction and anther browning rate, such as AgN03 concentration, active carbon and potato extract during the culture process were investigated using five genotypes of Ne16, Kexin13, VA, and Neishu7and Luyin 1. Callus of Ne16, Kexin13 and Neishu7 for experiment materials, four kinds of medium and three genotypes were complex variance analyzed. Ploidy level of the regenerated plants was determined by chromosome counting and the chloroplast number in stomatal guard cell.Access to the double haploid Micro-tuber transplanting and sowing, and hybridization, and identification of agronomic traits.The results showed that:1.The size of flower bud and size and color of anther could be used as simple but reliable index for judgment of the developmental stage of microspores. When the microspore characteristics of bud is in mononuclear margination stage, flower bud size is 5.0～6.0 mm ,the color of anther is light green.2.Cultivate in the same conditions, the different genotypes to anther culture, the performance of genotype differences in callus induction rate difference was observed.The frequency of induction was 8.1%. But the frequency of induction anther callus from Hu H9601-29 was 0.2%.3.The concentration of plant hormones and the hormone combination on anther culture play a decisive role.MS in this study as the basic medium by adding different concentrations of NAA, 2, 4-D and KT, the results showed that: MS + NAA 0.5mg / L +2,4-D 0.5 mg / L + KT 0.5mg / L suitable for callus induction.4. Potato anther culture of low-temperature pretreatment, mannitol pretreatment and heat treatment can obviously improve potato callus induction rate.Experimental results show that: potato anther 4℃under the conditions of the first treatment 2d, re-use of mannitol pretreatment 2d. Then inoculated, after inoculation of high temperature treatment 33℃, 24h best deal. Mannitol concentration of 40g / L of the appropriate effect5. Medium add AgN03, activated carbon and potato extract can improve the rate of callus induction; add AgN03, activated carbon can reduce the browning rate. 20mg / L AgN03, 0.5g / L of activated carbon and 50g / L of potato extract was the best.6. Sucrose as a carbon source, the concentration of 6% potato callus induction rate. When sucrose and maltose as carbon source, the concentration of 6%, the maltose on callus induction was higher than that of sucrose7. With Ne16, Kexin13 and Neishu7 of callus for the test materials, four kinds of medium and three genotypes were complex variance analyzed, LSR with the new mediums and genotypes on group for multiple comparisons. The results showed that: varieties > medium > varieties x medium, indicating that the differences between varieties significantly higher than the medium and species x medium, the difference between the medium was significantly higher than the species x medium. Between the medium and the differences between varieties was significant.8.The use of counting the number of stoma chloroplast in guard cell can be identified chloroplast counting chromosome ploidy plants. The test results showed that: Ne16 of haploid touraine stomatal guard cell chloroplast number at 10 ~ 20, range in the tetraploid range of 20 ~ 30, the chloroplast of the number of less than 20, for the double haploid than less than 20 and 30 for the tetraploid.9.Transplanting double haploid and tetraploid plants were at growth potential, leaf color, and other characters on potato-based comparison, test results showed clear differences. And access to the double haploid mini-tuber sowing and harvest. |
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