| In the middle of October 2001, a gigantic variant of tobacco was found in the tobacco field in the county of Xunyang, Shaanxi Province. In comparison with the contemporary control plants, the variant expressed several dominant growth characters, such as 3.4 meters high, 17.5cm stem girth, 129 leaves. In addition, the variant plant has a good quality. After being baked, the leaves had loose organization, citrine with bright gloss and rich oil content. However, this gigantic plant was rooted out before maturing, no seeds were produced. Fortunately, there were a few remained axillary buds on the plant. In order to rescue and reserve this resource, the variant was cloned through tissue culture and cuttage with axillary buds. The morphological comparison, cytological observation, physiol-biochemical and molecular biology characterization were carried out for cloned variant plants.Through cuttage of axillary buds and leaf tissue culture, numerous regenerated plantlets had been produced and were transplanted in the field. After treating with Img/L IB A for 7days, the variant axillary buds were planted in the sands. The survival frequency of sand culture was 86.7%. The optimized experimental condition of tissue culture showed that on MS medium added with Img/L 6B A, the variant leaf segment can directly differentiate buds (100%). The stems of the variant can extend quickly on MS medium added with 3mg/L GA3. The rooting frequency of bud was 63.08% on the 1/2 MS medium added with 2mg/L EBA. The plantlets of over 90% survived after transplanted in the field.The morphological comparison showed that the cloned variant plants were quite different from the controls. The cloned variant plants were transplanted in the field in the spring of 2002. Even though it was drought in the summer of that year, the cloned plants still kept the characters of the original variant plant, such as, good growth vigor, taller and bigger and with deep green leaves. By the end of October, the cloned plants were over 2.70 meters high with more than 110 leaves. In the end of November, it became colder and colder, the cloned variant plants had no flowers. As for the control, the average height was normally 1.22 meters high with 32 leaves, and it appeared flowers after being planted in the field for 65 days or so. The leaves of the variant plants, transplanted in the greenhouse were narrower and greener than the control's. The variant plants in the greenhouse appeared flower buds in the middle of December 2002, flowered in March 2003, and set seeds by the end of April. About 90% of plump seeds could germinate.The cytological observation showed that the cloned variant plants were diploid (2n=48), the same as the controls. But the chloroplasts number of the guard cells of stoma was different between variants and the control. The variant plant had 12.5 chloroplasts on average, while the control had 9.5 chloroplasts.The content of chlorophyll a, b and total chlorophyll in the variant plants were higher than in the control. The content of the soluble protein in the variant was 1.18 times as much as in the controls. Peroxidase isozyme and Cytochrome-oxidase isozyme electrophoresis analysis indicated that there were certain differences between the variant and the control in expression of enzyme. The soluble protein electrophoresis showed that 4 bands in the variant disappeared, such as Pl(l 14.6KD), P2(103KD), P3(66.2KD) and P4(24KD).The variant DNA and the control's DNA were investigated by RAPD analysis. The result showed that 18 out of 40 arbitrary primers produced distinctive bands. The variant and the control RAPD results were 4 types of patterns. First, the variant and the control had the same bands without difference. Second, the variant amplified more bands than the control. Third, the variant amplified bands were less than the control. Fourth, the variant and the control had the same bands, but the position was different. These results showed that the variant and the control similar index was 0.612, and there were also some differen...
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