Research & Utilization Of Purity Identification Technique Of Hybrid Wheat

Wheat is one of the world's most important food crops. With the development of hybrid wheat, seed purity has become the key factor which restricting heterosis application in wheat. In this research, to develop a simple and effective method suitable to seed purity test of hybrid wheat, three wheat hybrids and their parents were analyzed by acid polyacrylamide gel electrophoresis (A-PAGE) and random amplified polymorphic DNA(RAPD) technology. The results as follows:1. The standard gliadin fingerprints of'xz1','xz3'and'xz5'were constructed by A-PAGE. All the parents had characteristic bands. The gliadin fingerprints of'xz1'and'xz3'represented complemental.'xz5'had a characteristic band which can discriminate hybrids from their parents. Purity testing results of mixed seeds indicated that this standard gliadin fingerprints database could be used to seed purity identification of hybrid wheat.2. A simple, convenient and timesaving DNA extraction method from single partial seed with embryo was established. It could be used to RAPD analasis and seed purity test.3. RAPD optimum reaction system that can be used to identify hybrid wheat seed purity was built. The PCR mixture of 20μL contained 1×PCR Buffer, 2.0 mmol/L Mg2+, 400 nmol/L random primer, 0.75 units Taq DNA polymerase, 30 ng genomic DNA, 200μmol/L deoxynucleoside triphosphate (dNTPs). The amplification program of 35 cycles consisted of 1 min at 94°C, 2 min at 37°C, and 3 min at 72°C. A final extension step at 72°C for 10 min concluded amplification.4. With optimum reaction system, 23 out of 120 primers appeared to be polymorphic in different material. Primer S164, with clear and steady bands, could produce both maternal-specific and paternal-specific DNA bands in'xz1'. It could be used to seed purity test of xz1. In'xz3'and its parents, primer S158 represented maternal type, primer S170 represented paternal type; In'xz5'and its parents, primer OPD-05 and OPE-05 represented maternal type, Primer OPB-19 and OPF-14 represented paternal type. 5. Seed purity of'xz1'was identified by A-PAGE and RAPD technology. The identification results were 97.00% and 96.97%. There was no obvious difference beteen the results. It indicated that A-PAGE and RAPD technology also could be used to seed purity test of'xz1'.