Transformation Of Common Wheat By Zinc Finger Gene TaCHP

Wheat(Triticuma aestivum L.) is one of the most important crops in the world. Over 1/3 of the world population live on it, so the yield of wheat, affecting people's lives significantly.Abiotic stress has obvious influence on crop's growth and development, especially salt stress. Salt stress could cause lower yield and poor quality of crops. Wheat is a glycophyte, which resistance is controlled by polygene, so it is really difficult to obtain a salt-resistant species by common methods. With the development of molecular biological techniques, genetic engineering has already been used for cultivating new species, which have tolerance to salt and high production under salt stress.Transcription factors are a kind of multiprotein complex, which can combine with the cis acting element region on the promoter and regulate the expression of many stress-related genes, such as UV, drought, salt, etc., inducing various physiological and biochemical changes. The family of Zinc finger protein is a class of transcription factors, which has many activities, including resistance to stress. The salt-tolerance TaCHP gene came from a new somatic hybrid introgression line Shanrong No.3 (SR3), which has been generated in our lab from hybridization of common wheat Jinan 177 (JN177) with Thinopyrum ponticum, a salt and drought tolerant grass. In this study, five genotypes of wheat cultivars were transformed by Agrobacterium tumefaciens to the basal portion of seedling. In another experiment, co-cultivation with A.tumefaciens method was used for introduction of exogenous TaCHP gene into immature embryo derived calli (HeSheng 3) of wheat.1. The survival of transformed plantlets by the basal portion of seedling was: Jimai 17(430), Jimai 22(552), Yangmai 15(240), Longmai 30(520), Zhoumai(408). After selection with solution containing 100mg/L Hyg, and PCR detection with the primers UTA and TF8S, it is showed that the transformation efficiency is related to the genotypes of wheat. 2. Using the primer of UTA and TF8S to detect the transgenic wheat, we found the positive rate is distinguishing for the different genetype wheat. The PCR result of T1 and T2 generation show the positive rate decreased gradually, it is caused by the effect of chimera and the foreign gene translated into the meristem cell, which develop to leaf.3. We cultivate the T2 generation positive wheat to T4 generation in the greenhouse, detect the positive plant with southern blotting and analysis the expression of TaCHP. T4 generation positive wheat lines were treated with 200mM NaCl for 0,3,6,12,24 hours severally. The result of RT-PCR demonstrate the expression of TaCHP is upregulated in root and leaf.4. After T4 generation positive wheat and the control plant were treated with 0, 100,200mM for 14 days, we assay the differences between their leaves on proline content, MDA content, the activity of POD and the activity of CAT. With the treatment of 100 and 200 mM NaCl, the proline content in the transgenic wheat were higer than the control plant, although the proline content were rised in both of them. The MDA content were both rised with the increasing of salinity, but it in the transgenic wheat was low, comparing with the control ones. Similarly, the activity of POD and CAT were higer in the transgenic wheat.