Genetic Effects Of Dominant Genic Male Sterility(DGMS79-399-3) And In Vitro Conservation Of The Homozygous DGMS Genotype Materials In Cabbage

The dominant genic male sterile DGMS 79-399-3 in cabbage (as follows abbreviating DGMS) has been successfully used in seed producion in China, but there still have some defects in practice. : (1) some materials are difficult to obtain the sensitive male sterile plants which occasionally produce a few pollens, so it is difficult to obtain homozygous DGMS plants (MsMs) ; (2) the environmental sensitive trait of DGMS materials needs to be farther studied; (3) the genetic effects of sterile trait needs to be studied; (4) the genetic stability of the homozygous DGMS materials and TuMV virus changes after longer in vitro conservation; (5) the technology of in vitro conservation and proliferation for some DGMS genotypes needs to be further improved. To conquer these obstacles, lots of DGMS cabbage as experimental materials, the following areas were studied: 1.The relationship between the fertility of DGMS materials and the environment factors; 2.The methods to induce and obtain homozygous DGMS materials; 3.The genetic effects of the DGMS cabbage; 4.The detection of the genetic stability and the TuMV virus change during in vitro conservation of the DGMS cabbage; 5.The methods of in vitro conservation and increasing proliferation efficiency. The results are as follows:1. It showed that the DGMS materials with different genetic background were divided into the stable type and the sensitive type. The sterile degrees of materials 820 129 365 are very stable, so they have a potential in practice. But for the sensitive DGMS materials, temperature (mainly average temperature) was considered as the main environment factor affecting the fertile expression. The effect of temperature on the fertile expression was different among the different environmental sensitive DGMS materials. The lowest critical average temperature which leaded to 100% male-sterility in material 806 was about 20℃about 19℃for materials 811 813 and 22-24℃for 818 respectively. The temperature sensitive phase of the fertile transformation was 1-3 weeks before flowering, the most sensitive phase was 12-18 days before flowering.2. To reduce the environmental temperature was a method to obtain sensitive sterile materials with a few pollens. The DGMS materials 374 did not produce a few pollens many years was the first time to get the fertile flowers having a few pollens under the plastic tunnel conditions of the about 12℃average temperature. 20 plants randomly selected from the test-cross progenies of the male sterile material 374 were used to identify their genotypes by AFLP technology , the result showed that there have 7 homozygous dominant male sterile plants in total from 20 plants.3. Genetic effect of the DGMS in cabbage was studied with two different genetic-analysis models(the method of Griffing II and joint analysis of six generations). The result indicated that the best fitted genetic model to affect male sterile traits of the DGMS in cabbage was one major additive - dominant gene plus additive - dominant - epistatic polygenes(D model),the heritability of major gene was 80.02%-94.12% and the heritability of polygenes were 0.14%- 10.94%. The dominant effect of major gene was dominantly higher than the additive effect of polygenes. These results notly signified the main gene of affecting fertile characteristic of the DGMS cabbage was the major dominant genetic gene. Namely, the dominant effect of major gene had the important effect, however the additive effect of major gene and the polygenes effect should be also valued .The analysis results of genetic effects noted that the homozygous DGMS material 523-16 and 603-28 had the higher positive effect value of the general combination ability, and the average of sterile degree of five male sterile lines (combinations) coming from them was 99.19% and 98.18% respectively, and the sterile degree of three lines among the five sterile lines was 100% in the whole flowering period, so these two homozygous DGMS materials had a potential in practice.4. DNA detection of material Z15 (namely 96Z15 98Z15 and 99Z15) in vitro conserved 10 8 and 7 respectively by AFLP showed that there were no DNA-level variation. The DNA metylation status of 96Z15, 98Z15, 99Z15 obtained in 2005 for conserving half year were tested by MASP.A11 the three materials had pattern changes of methylation,de novo methylation and demethylation, but the degree of DNA methylation decreased in total.The TuMV virus of homozygous dominant male sterility cabbage in vitro conservation was tested by RT-PCR. Results indicated that the infection rate of TuMV virus in the bodies of tissue culture seedlings has the relationship with the infection rate of materials themselves in the field and the virus occurring quantity of conservation years; the tissue culture seedlings after subculturing two times had no increase in TuMV virus infecting rate.The DGMS cabbage in vitro conservation had DNA metylation status changes in vitro conservation and TuMV virus existed in the tissue culture seedlings, which did not affect the growth in the field and botanic traits of tissue culture materials. The above results showed that at present our in vitro conservation procedure is viable to conserve DGMS cabbages.5. The methods of conservation of DGMS materials were studied under the temperature of 18-20℃,.The segment having single shoot was suitable for material 04Z521-12 conservation and 2-3 shoots for 03Z -01Z608-1 conservation respectively. Sealing bottles by ventilation film + one layer plastic film can reduce subculture times of the 04Z521-12.The best culture medium for 03Z-01Z608-1 and 04Z521-12 was MS+6-BA0.2mg/L+NAA0.1mg/L+mannitol 0.3%+Active Carbon 0.02%+sucrose 3% and MS+6-BA 0.2mg/L+NAA 0.1mg/L+sucrose 7%, respectively. The materials after conserving were transferred to fresh culture medium, then 98% segments recovered growth.6. The fast reproduction method was studied for three DGMS materials 04Z521-12,03Z-01Z608-1 and 04Z-01Z126-1 with different head types. The fast reproduction medium for 04Z521-12 was MS+ 6-BA0.5mg/L+NAA0.1mg/L+sucrose3%, pH6.1, MS+6-BA0.5mg/L+NAA0.1mg/L+sucrose 2.%, pH6.1 for 03Z-01Z608-1 and MS+6-BA0.5mg/L+NAA0.1mg/L+sucrose2% , pH6.0 for 04Z-01Z126-1 respectively; optimal sealed material is ventilation film. Different male sterility lines had different proliferation efficiency because of different genotypes under the same culture condition. Round-head type DGMS material 04Z521-12 had the highest proliferation efficiency, while pointed-head type DGMS material 03Z-01Z 608-1 had the lowest proliferation efficiency.