Research Of Cold Resistance Of Artemisias Phaerocephala And Its △12 Fatty Acid Desaturase (FAD2) Gene Transformation Of Tobacco

A12 Fatty acid desaturase (FAD2) is the key enzyme which catalyzes the introduction of double bonds at the Δl2-position into the hydrocarbon chain and thereby converts oleic acid to linoleic acid. The contents of linoleic acid and unsaturated fatty acids were regulated by FAD2. Artemisia sphaerocephala is an endemic plant in desert and semi-desert areas of China. Linoleic acid content in A. sphaerocephala seeds oil was the highest among all plant studied, accounting for 80% of the total oil content. Using A. sphaerocephala as material, physiological changes under low temperature stress were studied; AsFAD2 gene cloned from A. sphaerocephalawas transfered into tobacco NC89 by agrobacterium-mediated transformation, and the effect of AsFAD2 expression on linoleic acid, cold resistance and plant growth and development were studied. The study provided basic information for breeding new variety with characters of high quality and strong resistance.The main results are as follows:1. At 10℃, the growth, relative electric conductivity,and MDA content of sphaerocephala seedlings showed little difference; At 6℃ and 2℃, linoleic acid proportion in total fatty acid, content of proline and the soluble sugar, as protective substances to low temperature resistance increased markedly; At 6℃, relative electric conductivity and MDA content increased at the beginning and then decreased slowly, the seedlings showed a certain resistance to 6℃; Seedlings had been greatly damaged after growing at 2℃ for four days, with the characters of slow plant growth, leaf water deficit, a sharp increase in conductivity and MDA content.2. Expression vector PBI121-AsFAD2 was imported into Agrobacterium GV3101 cells with the freeze-thaw method. AsFAD2 gene was transfered imported into tobacco NC89 by agrobacterium-mediated method,97 kanamycin resistance tobacco plants were confirmed as positive transgenic plants by PCR method, and expression levels of transcription in transgenic tobacco were validated by RT-PCR. Content of linoleic acid and linolenic acid in transgenic tobaccos leaves was 28.9% higher than that of the control, content of stearic acid was 39.6% lower than that of the control.3. After treatment at 2℃ for 12 h and 192 h, malondialdehyde content of transgenic tobaccos showed no difference between treatments and controls, and significantly lower than wide type tobaccos, while the content of proline was significantly higher than that of wide type. Transgenic tobacco showed greater cold than wide type one.