The Physiological Study And APRT Gene Cloning Of Thermo-sensitive Male Sterility Of160S

In higher plant, the male sterility is a popular phenomenon. Brassica napus male sterility plays an important role in the plant hybridization, which has important utilitive property. But at the morphological and physiological areas, there are little reports on thermo-sensitive male sterility of Brassica napus.We analyzed the changes of the floral organ morphology, the soluble proteins and physiological character of the B. napus160S in the fertility transition stage. The results of the floral organ morphology between160S, YB, HC8and160S×HC8showed that the petal of160S×HC8is biggest and the petal of the160S is smallest. In the ratio of the anther and the flower filament, the ratio of the160S is smallest, the anthers of the31C, HC8and160S X HC8are very plump. These three strains have enough anthers and their anthers have bright color, stable fertility which always maintains in a high level. The anthers of the160S are smaller and the160S did not have enough anthers. The anther fertility of160S is approximately equal to the anther fertility of160S×HC8under low temperature, but the anther fertility decreased dramatically when the temperature increasing, which approaches the sterility.All organisms can produce a range of ROS (Reactive oxygen species) which include superoxide (O2-), hydrogen peroxide (H2O2) and the hydroxyl radical (-OH) during the normal metabolic process. Malondialdehyde (MDA) is a widely used marker of oxidative lipid injury whose concentration varies in response to biotic and abiotic stress. Besides the antioxidants such as ascorbate and glutathione, there still exist ROS-scavenging enzymes, such as SOD (superoxide dismutase), APX (ascorbate peroxidase) and CAT (catalase). We analyzed the antioxidant enzyme activity and MDA content of the160S anther and31C during the fertility transition stage. The results showed that although the MDA content of the160S anther increases slightly when the temperature is lower than27℃. The MDA content increased significantly under the33℃, on the other hand, the MDA content of31C changed slightly. When the temperature is lower than27℃, the SOD, CAT, POD and APX activity of160S anthers increased with the rises of the temperature, which is similar to the antioxidant enzyme activity of the31C anthers. But the antioxidant enzyme activity of the160S anthers decreased significantly and the antioxidant enzyme activity of the31C anthers is still high under33℃. The results of the study between160S and31C bud soluble proteins during four stages showed that the concentration of the flower bud protein between the two strains had no significant differences. But in the category of the protein, the result of the SDS-PAGE showed that the160S lacks four stripes compared with the fertile strain31C, which probably causes the sterility of the160S pollens.In plant, adenine phosphoribosyl transferase (APRT, EC2.4.2.7) is the primary enzyme that converts adenine into adenosine-5-monophosphate (AMP) in the salvage metabolism pathway. We cloned the adenine phosphoribosyltransferase (APRT) gene from the thermo-sensitive and male sterile Brassica napus160S. We obatained its partial gene sequence and its length is1400bp. From the result of blast in NCBI (National Center for Biotechnology Information), we could observe that it matched with the APRT CDS sequences of the Arabidopsis and Brassica napus (GenBank:AY324112.1) in the percentage of96and98.