Cloning And Fuction Study Of V-ATPase Subunit B Of Arabidopsis Thaliana

Vacuolar ATPase (V-ATPase) including 12 subunits is a multi-subunit complex which can be divided into two structural domains. V-ATPase is responsible for many important cell processes, such as acidification of intra-cellular, endocytosis mediated by receptor, post-translational translocation and membrane fusion of some organelles and plasmolemma and so on. Each subunit of V-ATPase is inseparable for the holoenzymes, however, there is hardly any research or report about the V-ATPase of higher plant especially model plant Arabidopisis thaliana.We first initial researched the fuctions of B subunit of V-ATPase in Arabidopisis thaliana. There are three isoforms of V-ATPase, VHA-B1, VHA-B2 and VHA-B3.1 The tissue-specific expression of V-ATPase B subunit were displayed by the GUS reportor gene. They express in various organs, including whole seedling, root, flowers, pollen grains and pollen tubes after germination for 4h in vitro.2 For function studying of V-ATPase B subunit of Arabidopisis thaliana in vivo, we have overexpressed the V-ATPase B subunit and choosed T-DNA insertion mutant. The overexpression vector fused of yellow fluorescin and started expression by CaMV35s or pollen special promoter lat52 were constructed and then transgenic plants expressing Fusions VHA-B subunit were choosed through hygromycin and fluorescence detection. These overexpression plants could be used to reveal the contribution of VHA-B subunit to growth, development and reproduction of Arabidopisis thaliana.3 T-DNA insertion mutant is the effective tool to research gene fuction of plants. We have identified the T-DNA insertion mutants about VHA-B1, VHA-B2 and VHA-B3. Some of them were homozygote which will be verified at transcription and translational level, so as to observe the phenotype of mutants. The heterozygote will be continued to grow until isolate the homozygous.This subject revealed the tissues and developmental special expression of V-ATPase B subunit promoters as well as the abnormal phenotype when overexpression or missing VHA-B subunit in Arabidopisis thaliana. With understanding the molecular mechanism of VHA-B subunit, there will be more bases for the study of V-ATPase.