| This study uses P. forbesii, which is endemic to China as test materials, and holds a systematic research of its characteristics of introducing cultivation, and its biological characteristics of flowing, pollination, fruiting. And the research also analysis the karyotype of five primroses genera plants P.forbesii, P.maximowiczii, P.serratifolia, P.sinensis, P.secundiflora Franch.Establish the rapid propagation system of the group growing P.forbesii and use the colchicine to polyploidy induce the P.forbesii. Using morphological and anatomical,cytology,DNA content,chlorophyll fluorescence analysis and such kind of means to conduct polyploid appraisal the purpose is to induce a polyploid P. forbesii to improve its appreciation,enhance its resistance.With its domesticated success and cultivating new polyploid primroses varieties, to make full use of our country's wild resources advantage, and have important meanings in the protection of resources, develop cultivation in China. Main results are as follows:1,The results of the Biological characteristics of P forbesii study indicate that: The fruit of P.forbesii is capsule, spherical, after dehydration the epicarp will craze membranously when mature, outside have 1-nerved calyx. Normally mature seeds are tiny, irregular hexahedral, their epidermis have mesh shape structure. The seeds of 1000-grain weight (0.118±0.0023) g. The seeds of the P.forbesii should be stored for dry at low temperature, if storage at the room temperature, after a year most seeds lose their vitality. GA3 in improving seed germination rate effect is not big, but the GA3 treatment of P.forbesii seed germination time is short, sprout neatly. The seeds of P.forbesii is suitable to make a spring at a germination temperature 20 ~ 25℃. P.forbesii is lean to weakly acidic water and soil environment. So in domesticated P.forbesii, the soil and the water should be improved to pH = 6.8 in order to favor the seed germination, make it normally grow up. In A. budding soil corrosion: peat moss: sand = 2:1:1 as train matrix helpful for P.forbesii's seed germination, germinating capacities and sprout potential is obviously higher than other matrix. So in planting early introduction cultivation matrix should choose (budding soil corrosion: peat moss: sand = 2:1:1) as reproductive growth of soil matrix.P.forbesii are typical different flowers pollination plants.The bloom days of P.forbesii in Yunnan are from around November 15 to March 26, without night blossom. During Sunny days, the blossom peak ranges between 11:15~11:45. During the Cloudy days, the blossom hours are more scattered than during the sunny days, and flowering peak backward delayed. Through the determination of the pollen life force of P.forbesii, the pollen could be cultivated in 25℃, which has the highest rate, within 30min have pollen germination, cultivate 120min after, pollen germination rate can reach 48.92%, after 180min observation shows pollen germination rate can reach 62.87%, the influence of temperature and sucrose for P.forbesii pollen germination shows that the optimum temperature conditions is 25℃, sucrose concentration of 10%.When boric acid concentration is 10mg/L, P.forbesii pollen germination rate is the highest. P.forbesii has the best storage result when stored in low-temperature -20℃add desiccant. From the body pollen germination method is suitable for the life force test of P.forbesii. P.forbesii belongs to 3-fitting hole groove pollen. The long and short flower of P.forbesii has very different the pollen grains sizes, short styles of the pollen grains is larger, but long styles of the pollen grains is lesser, long styles have longer stigma and papillary cells, long styles papilloma covenant is of 1.5 times of the length of short styles, the long styles cells obviously longer than short styles cells. The Hybrid seed setting of Long styles flowers (♀)×short styles flowers (♂) or short styles (♀)×long styles flowers (♂) combinations are higher.2,The study of 5 kinds of primroses genus karyotype indicates: P.forbesii,Somatic chromosome number is 2n=2x=18, Brachial index is 51.65%, Karyotype formula is K(2n)=2x=18m, The karyotype asymmetry genera 1A. Chromosome of P.secundiflora Franch is larger, Centromere is more clear. Somatic chromosome number is 2n=2x=22, Brachial index is 52.00%, Karyotype formula is K(2n)=2x=20m+2sm, The karyotype asymmetry genera 1A. P.sinensis Somatic chromosome number is 2n=2x=22, Brachial index is 53.42%, Karyotype formula is K(2n)=2x=20m+2M, The karyotype asymmetry genera 1B. P.maximowiczii,Somatic chromosome number is 2n=2x=22, Brachial index is 51.07%, Karyotype formula is K(2n)=2x=22m, The karyotype asymmetry genera 1A.P.serratifolia Brachial index is 52.59%, Karyotype formula is K(2n)=2x=22m, The karyotype asymmetry genera 1A. except the P.secundiflora Franch, the karyotypes of other four plants are first reported.3,The result of the building of P.forbesii tissue regeneration system and rapid propagation autopolyploid induction system shows that use blades of callus induction formed fast, texture closely. Alexipharmic disinfection mode selection method of dealing with Numbers is 8, namely: detergent cleaning 20min, tap water flushing 30min, ShengGong soak, 7min 0.1% sterile water flushing 4~6 times. Hormone concentration ratio in olumes is 0.5 mg·L-1, 6-BA concentration ratio is 3.0 mg·L-1 when callus induction 85.0%, and the highest for the callus forms faster, texture closely in peak green, which has differentiated ability. The most suitable hormone formula of multiple shoot clumps induction medium for P.forbesii is MS + NAA0.10 mg·L-1 + 6-BA1.0 mg·L-1 (pH = 5.8). With 1/2MS for basic medium, NAA concentration would be 1.00 mg·L-1, which can produced the most root in the shortest possible time, rooting rate is 80.0%, and roots are very strong.It is an effective method in induction of polyploidy ornamental plants to use colchicine processing the clump sprouts during the process of tissue culture produces.With the 0.10% colchicine to process the clump sprouts of P. forbesiifor 48 h, which has the best inducing effect, the mortality rate is 15.0%, mutation rate is 70.0%, which is the best treatment combinations. Cell chromosome identification of the variant plants eventually results in strains of autotetraploid strains, with 13 strains from colchicine on clumps shoot induction and 6 strains are from colchicine on callus induction. The combination of colchicine and tissue growing skills will get much more polyploidy plants than in the field when using the polyploidy skills. In the field, there could not have induced tetraploid plants4,The study of morphology and cytology on P.forbesiidiploid and tetraploid shows that: tetraploid P.forbesii's heights are more shorter, petioles are more shorter and thicker, leaves are more wide and thick, its color is dark green, and more longer hair in the back of the leaves than diploid P.forbesii. Tetraploid plants, the average leaf index is 1.09±0.07 in contrast with 1.38±0.24 of the diploid average leaf index, tetraploid single corolla diameter increased significantly with an average of (2.27±0.03)cm, in contrast with(1.24±0.55)cm of the average corolla diameter, and the flower sepals, Yellow eye increased as compared with the control. The fruits are Larger than the ones in control, the fruit of tetraploid has an average diameter of(0.89±0.07)cm, whereas the fruit diploid has an average diameter of(0.41±0.15)cm, the difference was extremely significant. Stomatal guard cells of tetraploid plants were significantly greater than the diploid in the length and width, diploid guard cell chloroplasts is an average of 18, the average number of tetraploid guard cell chloroplasts is 55, reached a significant difference. Stomatal, Guard cell size and chloroplast's number and ploidy is a significant positive correlation. Diploid stomatal density is(94.20±8.56)/mm2, tetraploid stomatal density is(78.91±6.97)/mm2, the stomatal density on the existence of diploid and tetraploid are significantly different. By chromosome analysis of colchicine processed cell regeneration plants, we found that some of the regenerated plants are tetraploid (2n = 4x = 36). By karyotype analysis, tetraploid somatic of metaphase chromosomes consist of 36 metacentric Centromere chromosomes. Brachial index is (51.32±0.76)%, karyotype formula is K (2n) = 4x = 36m, the karyotype asymmetry is 1A. To further determine the doubling plants homozygous of ploidy, using flow cytometry and PartecDPAC to conduct diploid and part of plants which were chromosomes identified 2n = 4x = 36, measured and analyzed cellular DNA content. he results show that the leaves of the chromosome doubling plant, whose DNA content in cells in a corresponding increase doubled, which is consistent with the identification chromosome numbers.5,The preliminary study results of the chlorophyll fluorescence of P.forbesii diploid and tetraploid show that: Tetraploid P.forbesii's chlorophyll a, chlorophyll b, total chlorophyll and carotenoids are higher than diploid's. In diploid and tetraploid, the tetraploid's Fo is smaller than diploid, but Fm, Fv, Fv / Fm, Fv / Fo and PI is higher than diploid, which indicates that tetraploid have better photosynthetic performance. |
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