| Soil salinization is one of the main environmental stress factors that affect crop production in the world today. It is one of the most effective and feasible way to develop and utilize large area saline alkali land by using plant salt tolerance genetic engineering. Gene cloning and identification of salt tolerance in crops is the precondition of crop salt tolerance genetic engineering. Oat with drought tolerance, salt resistance and barren resistance and other characteristics, is an important genetic resource for crop improvement of resistance. It is very important to study the function of oat salt tolerance gene by using the mining and utilization of the salt tolerance gene. The purpose of this study is to clone the salt resistant genes of oat based on the previous work, and to verify the salt tolerance function. The main results are as follows:1. Using RACE technique, a KUP/HAK/KT family gene was cloned from oats, which was named AsKUP 1. The gene cDNA full-length 2951bp, encoding 777 amino acids. isoelectric point is 8.55, molecular weight is 87011.80. GenBank accession number is KX034554. Prediction of AsKUP 1 transmembrane domain, the results show that the protein has 14 transmembrane domains, which indicates that the protein is a transmembrane protein. Prediction of AsKUP 1 protein subcellular location, the results show that the cell membrane is located in a greater probability. Predicted protein Pro/ hydrophobicity, as a result of hydrophobic proteins. Using PCR technology, a NAC family gene was cloned from oats, named AsNAC1. The gene cDNA full-length 1710bp, encoding 320 amino acids, isoelectric point is 6.62, molecular weight is 35454.99. GenBank accession number is KU886332. Prediction of transmembrane domain of AsNACl, the results showed that the protein had no transmembrane domain. Prediction of AsKUP 1 protein subcellular location, the results show that the greater the probability of nuclear. Predicted protein Pro/hydrophobicity, and the results were shown as hydrophilic proteins.2.The full-length coding sequence AsKUP 1 gene and AsNAC1 gene of the successfully constructed the plasmid pCAMBIA1301 plant expression vector, using inflorescence infection was transformed into wild-type Arabidopsis thaliana, using hygromycin antibiotic resistance screening, and detected by PCR amplification, were obtained 3 turn tree gene AsKUP I positive plants and 5 tree gene conversion AsNAC1 positive plants. Through the screening of Mendel’s law, the transgenic AsKUP1 lines had 2 strains, which were close to 3:1, and the transgenic AsNACl lines had 3 strains, which were close to 3:1.3. In medium containing 150 mmol/L NaCl MS, turn AsKUP1 Arabidopsis seeds and turn AsNAC1 Arabidopsis seed germination rate than wild-type Arabidopsis seed germination rate is high; turn AsKUP1 Arabidopsis lines line 1 and line 2 were wild type plants root length of 1.46 times and 1.34 times, significant difference; turn AsKUP1 Arabidopsis wild type plant fresh and dry weight of 1.56 and 1.44 times, significant Mfference; proof of AsKUP1 had increased expression of the Arabidopsis salt tolerance. Turn AsKUP1 Arabidopsis K+ content was significantly higher than that in wild type Arabidopsis K+content, and the content of Na+ without significant difference, the conjecture turn AsKUP1 genes in Arabidopsis thaliana plants is by absorbing K maintain higher K+/Na+ ratio to improve the salt tolerance. Turn AsNAC1 Arabidopsis lines lines 1, strains 2 and 3 were wild type plants root length 1.39 times,1.52 and 1.37 times, significant difference; turn AsKUP1 Arabidopsis wild type plant fresh and dry weight of 1.50 times and 1.49 times, significant difference; proof of AsNAC1 had increased expression of the Arabidopsis plant salt tolerance. |
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