Study of Glutamine Synthetase Overexpression Mutant in Arabidopsis

• Main Authors: Chia-Chien Wu, 吳佳建
• Other Authors: Tsung-Luo Jinn
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/04768454356308690064

碩士 === 國立臺灣大學 === 植物科學研究所 === 92 === Leaves are the most noticeable plant organs. They are predominant photosynthetic organs of a plant and pivotal for carbon fixation. However, less is known about the molecular genetic interactions of leaf morphogenesis. Our laboratory screened for morphological mutants of Arabidopsis thaliana harboring T-DNA cassette from pPZP202:BAR:SK, a T-DNA tagging vector, with the purpose of contributing to the dissection of the mechanisms underly leaf ontogeny. Here, we showed a gain-of-function mutant, vig1-D, with broad, rounded lamina and conspicuous venation phenotypes. Southern blot and inversed PCR identified two T-DNA linked together and inserted in the downstream of At5g16570 (VIG1), encoding a putative glutamine synthetase (GS). The amino acid sequence alignment indicated that this protein contains two important conserved domains and shows highly similarity, above 79%, to other GS1 members in Arabidopsis. Using Northern blot and RT-PCR, we showed that the transcripts of VIG1 were overexpressed in vig1-D. Moreover, the expression of other related GS genes, such as GLN1;1, GLN1;2, GLN1;3 and GLN2, was also increased in the stems or leaves. Constitutive overexpression of VIG1 in wild-type plant recapitulated the vig1-D phenotype, confirmed that increased expression of VIG1 was responsible for the abnormal phenotype of vig1-D. In previous studies, many evidences have indicated that transgenic plants overexpressing the GS also showed that phenotypes such as early leaf senescence were similar to vig1-D. PVIG1:GUS analysis in transgenic Arabidopsis showed that VIG1 was expressed specifically in leaf trichomes, thus suggesting altered morphology of vig1-D due to the ectopic expression of VIG1 in leaves. Therefore, these results suggested that regulation of the ammonia homeostasis in plant may be involved in the leaf morphogenesis.