| Tobacco (Nicotiana tabacum L.) is an important economic crop and also one of the important pillars of the national economy. At present, the parents for tobacco breeding are relatively concentrated and germplasm genetic base is relatively narrow. It is difficult to produce new varieties of tobacco. Therefore, it is necessary to examine germplasm resources' genetic diversity among the tobaccos for providing a theoretical basis for parents choice of the tobacco breeding and the maximum utilization of tobacco germplasm resources.Nitrogen is one of three major nutrition elements and also one of the rate-limiting factors in tabacoo growth. Cost of nitrogen fertilization accounts for 40% of the total cost in tobacco production. Transforming ammonium into the glutamine catalyzed by Glutamine synthetase (GS) is an important step in tabacoo nitrogen metabolisim pathway. It is also a key limited-rate procedure in nitrogen assimilation. The nitrogen utilization for plant needs the participation of a large amounts of carbon skeleton (a-ketoglutarate). The level of carbon plays a positive role in GS expression. The transcription factor Dof1 is closely relation to expression of relative carbon metabolism gene. Present research shows that traditional breeding method is difficult to breed crop varieties with enhancing nitrogen utilization. So it is expected to over-express GS and Dof1 which related to carbon metabolic pathway in tobacco by using plant genetic engineering technology to improve nitrogen utilization and the tolerance to low-nitrogen stress of tobacco. This paper obtained the following results:1. The genetic diversity was assessed among 25 tobacco accessions by ISSR marker, and five effective primers were screened out of the 50 ISSR primers. Those five primers finally amplified 100 allels, of which 76 allels were polymorphic, and the ratio of polymorphic allels was 76%. The Jaccard's genetic similarity (GS) coefficients were calculated by NTSYSpc-2.10e software. The result indicated that the GS coefficients among the 25 tobacco accessions were ranged from 0.313 to 0.938 with an average of 0.737. The GS coefficients between three wild tobacco accessions and the cultivated tobacco accessions were smaller than those among the cultivated tobacco accessions, and this suggested that the genetic variation was high between the wild and the cultivated tobacco accessions. The 25 tobacco accessions were divided into two groups on the UPGMA dendrogram constructed from GS coefficients. The three wild tobacco accessions were clustered into one group, while the 22 cultivated tobacco accessions fell into the other group.2. GS1 and GS2 were separated from Arabidopisis cDNA by Polymerase Chain Reaction (PCR). The constitutive/inducible plant expression vectors pH2-35S-PRbcS-GS1 and pK2-35S-PRbcS-*T-GS2 which controled by CaMV35S and RbcS had been constructed.3. These constitutive/inducible plant expression vectors pH2-35S-PRbcS-GS1 and pK2-35S-PRbcS-*T-GS2 were respectively transformed into wild type tobacco (WT) and the transgenic tobacco which had been transformed plant expression vector pPZP221-PRbcS-Dof by Agrobacterium-mediated method, we got six kinds of transgenic tobaccos: WT-GS1 , WT-GS2 , WT-Dof1-GS1 , WT-Dof1-GS2 , WT-GS1-GS2 , WT-Dof1-GS1-GS2. These transgenic tobaccos were confirmed through antibiotics selection and test of genomic PCR and RT-PCR.4. The datas of GS activities in 7 kinds transgenic tobaccos showed that the transgenic tobaccos' GS activities were 1.44-2.75 folds compared with control plants (WT). Determine the seedling height of transgenic tobaccos WT-GS2, WT-Dof1, WT-Dof1-GS1, WT-GS1-GS2, WT-Dof1-GS1-GS2 and wild-type tobacco cultivated under the condition of low nitrogen stress, the ratios were: 1.14, 1.23, 1.71, 1.43, 2.27, respectively. These showed that overexpression of GS1, GS2 and Dof1 can improve the GS activities and promote transgenic tobaccos' growth under the condition of low-nitrogen.
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