Analysis of the mechanism of tracheary-element differentiation using gene transfer techniques

1. 气管元件分化特异性表达基因的分离与鉴定：从百日草叶肉细胞中分离气管元件分化细胞MRNA，培养48 h，构建CDNA文库，并在诱导分化和非诱导分化培养基中对细胞MRNA进行差异筛选。结果，我们成功分离出4个在分化细胞中优先表达的基因（TED 1-4）。这些基因在培养48 h后迅速表达。这些基因的核酸序列表明，ted1与线粒体fl - atp酶对应，TED2与豚鼠晶状体蛋白相似，ted4与糊粉蛋白特异性蛋白相似。基因转移系统的建立与转移。利用农杆菌构建拟南芥和百日草的正义和反义DNA转移体系。将TED2的正义和反义DNA导入百日草细胞。将转化后的愈伤组织用于检测导入DNA对管状细胞分化的影响。然而，引入的dna对百日菊愈伤组织管状细胞的分化没有影响。

1. Isolation and characterization of genes expressed specifically to tracheary-element differentiation :A CDNA library was constructed from MRNA derived from cells differentiating to tracheary elements, which were isolated from Zinnia mesophyll cells and cultured for 48 h. A differential screening was performed in the library with MRNA from the cells cultured in differentiation-inducing and non-inducing medium. As a result, we succeeded in the isolation of 4 genes(TED 1-4)expressed preferentially in differentiating cells. These genes were expressed rapidly after 48 h of culture. The nucleic acid sequences of these genes indicated that TED 1 corresponded to mitochondrial Fl-ATPase, TED2 was similar to guinea pig lens protein and TED 4 was similar to aleurone specific protein.2. Establishment of gene transfer systems and the transfer of. sense and anti-sense DNA of TEDs into Zinnia cells :Gene transfer systems of Arabidopsis and Zinnia were established with Agrobacterium tumefaciens. Sense and anti-sense DNA of TED2 were introduced into Zinnia cells. The transformed callus was used for the examination of effects of introduced DNA on tracheary element differentiation. However, the introduced DNAs gave no effect on tracheary element differentiation in Zinnia callus.