Analysis Of The Gene Expression Related To Panicle, Anther And Taigu Genic Male-sterile In Wheat

Male sterility of wheat is important to heterosis utilization. DDRT-PCR is a method to analyse and clone differentially expressed gene and is widely used in plant. The work analyzed genes' expression of panicle, anther and Tal of near-isogenic-lines of Lul5 by DDRT-PCR. The main results were as follows:1. The total RNA was reverse transcripted by three anchor primers respectively and then was amplified by 30 single random primers. We have identified, out of a total of about 1,000 bands, 102 fragments where the expression of the corresponding genes seemed to expressed differentially, which involves fertility, sterility and stage specific genes. We selected 30 repeatable bands to be transfered to membrane. One of which were proved to be positive by RNA rev-Northern hybridization using [ a -32P]dCTP labeled first strand cDNA as probes. It was named as SI 1766ooand expressed increasingly in anther.2. Double primers , single RAPD primer+anchor primer and correspond single primer were used to PCR amplification. The result showed that :there were great difference in the banding patterns of the display among the three primer combinations: the bands of using double primer were fewer than those of corresponding single primer, furthermore, most of them were different; there were also great difference in those of the various single primers; the banding patterns of the single primer+anchor primer combination were similar to those of corresponding single primer. So it was proved that most of the bands were the product of the single primer, and we can select primers amplified stablely used for differential display effectively.3. Compared to EB-staining and agarose gel electrophoresis , silver staining and non-denature PAGE had many advantages such as higher resovling power and sensitivity, non-EB-pollution and the gels can be conserved longer. But the procedure was more complicated and more time was needed; Compared to auto-radiography and denaturing PAGE, it had also many advantages such as simpler procedure, less expense and non-radioisotope-pollution, but it has lower sensitivity. In a word, this detection method is the most practicable one. And the silver staining method of Sanguinetti is effective in detection of PCR products, which stains rapidly and has stable results.