| Two commercial cotton varieties, Xuzhou 142 (upland cotton) and PimaS-4 (sea island cotton) with normal fibers, were used as controls and compared with three fiber mutants, Xuzhou 142-fl without lint and fuzz (lintless-fuzzless), Xuzhou 142-N without fuzz (fuzzless) and Liong lintless with very short fibers (only about 6~8mm length), to investigate some important changes in expression of genes, activity of enzymes, biosynthesis of cellulose, sugar and malate during the differentiation and development of cotton fiber cells. The results evolved several aspects as follows:1. Comparison of extraction RNA methods from the cotton fibersThe comparison of three methods of RNA isolation with each other was conducted to isolate high-quality RNA from cotton fiber cells. The analysis show that the method by using an extraction buffer with high ion-density and pH value was better. This extraction buffer with high ion-density and ph value was characterized by adding extra reagents include sodium deoxycholate and Nonider-40 (NP-40), enhancing density of other salt compounds, elevating the value of pH, and supplementing polyvinylpyrrolidone (PVP-40) and 2-mercaptoethanol. High quality RNA from cotton fiber and other tissue cells were obtained by using this isolation method, for example, the fiber cell RNA can reach 1.85 of the 260/280 ratio with the yield of 0.65 mg.g'1 and the cotton leaf RNA 2.02 with 6.35 mg.g'1. It was this method that RNA isolated from cotton tissues, especially from fiber cells, were suitable for the experiments of RT-PCR and Northern blot hybridization.2. The effects of Sucrose synthase (SuSy) on the fiber initiation, elongation and thecellulose biosynthesisWhen comparison of the SuSy activity at ODPA (day post anthesis) with that at -1DPA in five different types of cotton ovules, it was found that ovules of Xuzhoul42, PimaS-4 and Ligon lintless have higher activity at 1DPA than at ODPA, increasing 169.05%, 154.84%, and 68.11%. However, in the lintless-fuzzless or fuzzless mutant, the enzyme activity declined from 51.4% to 64.7%. In other hand, sucrose in the ovules of fuzzless-lintless mutant had a higher content than that in ovules with fiber or fuzz during initiation of fibers from ODPA to 1DPA. It was suggested that the SuSy activity in ovules in the anthesis day would pay an important role in the fiber cell initiation.From 10DPA to 30DPA, the SuSy activity kept higher in fiber cells than embryo and seed coat cells. Except Ligon-lintless cotton fiber, the SuSy activity in all of other materials reached the peak value at 20 DPA, which coincided with the onset of cellulose synthesis of the secondary cell wall. The activity peak in Ligon-lintless cotton fiber was early occurred at 15DPA. After that, the SuSy activity in fiber and seed coat cell remained relatively high. So SuSy was more essential for cellulose biosynthesis than fiberelongation.3. Sucrase would regulate the fiber cell elongationFrom -1DPA to 5DPA, there was no sucrase activity in ovule of all cotton lines in the experiment. During 10-40DPA, die sucrase activity in Xuzhou 142 fiber cell obtained the highest at 15DPA, and then dropped rapidly, which was in accordance with the fiber cell rapid elongation. PimaS-4 (Sea island cotton) fiber cell had the peak at 20DPA. But the level of sucrase activity in Ligon-Lintless fiber cell was lower than Xuzhou 142 and PimaS-4, and also had no peak value. In the Lintless-Fuzzless mutant ovule, we almost haven't detected the sucrase activity. Surprisely, the activity of sucrase in Xuzhou 142 ovule was also undetectable at 10DPA. So we conferred that there are somethings which may inhabit the sucrase activity in the embryo or seed coat cells.4. The effects of soluble sugar and matale content on the fiber initiation, fiber elongation and cellulose biosynthesisThe soluble sugar content had no obvious change in five material ovules during -1-2DPA, but when compared with that at -1DPA, the soluble sugar content at 5DPA had a sharp rise, especially in XuZhou 142, sea isl... |
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