The Study Of Dongnong 303 Potato Introduced With Gene P5CS By Agrobacterium Mediated Method

Drought, salinity, heat, cold and other environment conditions are the important factors of affecting plant growth and development. These non-biological stress factors will affect the plant cell turgor pressure and cause the plant poor growth by osmotic stress. Under osmotic stress, the plant will produce osmotic adjusting substances, include proline and betaine, to resist osmotic stress. The proline is mainly derived from glutamate synthesis andΔ1-pyrroline-5-carboxylate synthetase(P5CS) is the key enzyme in proline synthesis. In this paper, we had transformed the P5CS gene into Dongnong303 potato genome in order to improve the osmotic stress of potato cultivars.First of all, the high-redifferentiation system of Dongnong303 potato was optimized by testing different hormone combinations. And then, the P5CS gene was transferred into potato genome by Agrobacterium-mediated method. At last, the insertion of target gene was verified using the methods of PCR, BLAST and Southern blot. In addition, the changes of transgenic plants growth and physiological were discussed under drought and salt stress.1. Virus-free potato plantlets and tubers were used to do transformation testing by different IAA, IBA, ZT and hygromycin combinations. The optimal differentiation condition for tubers was MS base medium complemented 3.5mg/L ZT+1.0mg/L IAA and hygromycin stress was 6mg/L. The best medium for rooting was 1/2 MS medium and selection pressure was 10mg/L.2. Molecular biology testing was taken on the plants selected by hygromycin. Gene-specific PCR using specific primers obtained the required band of 701bp. The band was BLAST with the P5CS gene of Arabidopsis thaliana after linking, transforming and sequencing. The result showed that the homology was 100%. And then by further southern blot, we have got one positive band suggesting that in the potato genome there was one copy of P5CS gene. We eventually obtained one transgenic lines.3. The changes of growth and physiological in transgenic plants under salt(1.5% NaCl) and drought(60%,40%,20%) were examined. Results showed that transgenic plants had much advantage than non-transgenic in range of extension of leaves, roots and quantity of tubers. In the matter of physiological and biochemical, the content of proline in transgenic plants increased more than 25 times than that in untreated ones, and it was 1.7 times of that in non-transgenic plants treated in the same way. At the same time, the content of MDA and activity of SOD were respectively 0.4 times and 2.6 times of that in non-transgenic plants. Under drought treatments of 60%,40%, and 20%, the content of proline in transgenic plants were respectively 1.2 times,1.4 times and 2.1 times of that in plants which was not transformed. Increase of MDA content in transgenic plants was 0.6 times of that in non-transgenic plants. And, activity of SOD in transformant was respectively 2.5,2.8, and 5.6 times of that in non-transgenic control. These results showed that potato transforming by P5CS gene could improve the stress to salt and drought tolerance.