Cloning, Subcellular Localization And Functional Analysis Of Sec27p,ARHI, Two Of Up-regulated Gene Group Related Vesicle Trafficking Response To Iron Deficiency In Rice

As dominating foodstuff, iron malnutrition forms one of the most serious problems for human health due to the relative iron-inefficiency of rice. With accomplishment of the rice genomic sequencing, function genomics research became the most effective tool in strategy studies on expression and regulation of rice genes, responded to Fe-deficiency.In previous research, our lab had acquired a group of the hightest -Fe/EDTA-Fe ratio genes related vesicle trafficking response to iron deficiency in rice based on the Microarray analysis. In this thesis, further research of the genes was performed.Observing ultrathin sections in root tip cell of rice through transmission electron microscopy(TEM), found protuberant vesicles beside plasma membrane, the vesicles in root tip cell of iron deficiency is bigger and more inclusion than ones of iron supply, and the vesicles in root tip cell of iron deficiency generated from cytoplasm to extracellular and diverse direction, the vesicles in root tip cell of iron supply only generated from cytoplasm to extracellular.16 up-regulated genes related vesicle trafficking response to iron deficiency in rice were blasted and analysed. Four genes, ARHL Sec27p, Tuftelin-interacting, pV72, were examined through RT-PCR and REAL-TIME PCR.Full length cDNA sequencesof Sec27p and ARHI were acquired and plant expression vectors pCAMBIA1302-Sec27p, pCAMBIA1302-ARHI were constructed respectively, recombinant plant expression vectors have been transformed into onion epidermic cells, then localizated object gene through observation of GFP instant expression. The results indicated that Sec27p was localized in the vesicles near to cell membrane, ARHI localized in nucleus and nearby membrane structure. Meanwhile, plant expression vectors had been transformed into tobacco using leaf disc transformation. Putative function analysis was carried out by NCBI sequence blast and sofeware of protein functional forecast. At last, Sec27p can be preliminary considered as a coatomer subunit involved in endoplasmic-to-Golgi protein trafficking. It is a powerful evidence of vesicle trafficking enhancement in root cells response to iron lacking. ARHI can be considered as a novle Small G-protein, regulates genes expression by modulating protein and RNA transport through nuclear pore complex.