The Labelling Of CesA1and ABP1and Their Co-Location Analysis

Auxin binding protein1(ABP1) is a new recognized receptor in auxin signal pathway. The ABP1mediates cell expansion and involved in the response of auxin signal to induce of the cell elongation. As we know the plant cell elongation is an actually facilitate process of accelerated vesicular transportation and activation of cell wall synthesis. In order to do a further study of the auxin signalling that the molecular labeling of the ABP1and cellulose synthase are carried out and transformed in this paper.Cellulose synthase1is a key enzyme in Cellulose synthesis of primary cell wall. In this study the two chimera expression vectors are constructed to separately label ABP1and CesA1with fluorescent protein respectively. We first cloned the cDNA of ABP1and CesAl from tobacco. Then the cDNA were constructed into the chimera structure that the ABP1with GFP and CesA1with CFP. The two recombinants are transformed into tobacco leaf for the transient expression in the same epidermal cell via infiltration to verify the construction. The confocal observation shows the two colors fluorescence is in some identical pavement cell thus the ABP1and CesA1proteins are successfully labeled. After scan the infiltrated leaf disc with the laser that the green and cyan florescent are separately observed and recorded in the same pavement cells. The pictures are then overlaid and the overlaid colors are analysis. The strong colors are accumulated in the cell membranes that mostly are overlaying especially in the lobe of the pavement cell. It demonstrates that the two proteins are distributed in the membrane with same manner implies the maybe effecting each other.The further Agrobacterium tumefaciens mediated transformation is applied to transfer the two recombinants into the model cell line BY-2. The two transgenic cell lines are screened out. It is observed that the ABP1-GFP transgenic cell is growing faster than the wild type. The transgenic CesA1-CFP calli spread out faster than its wild control. The two recombinants were transformed into the same BY-2cell by repeat Agrobacterium tumefaciens LBA4404co-culture transformation. The binary transformed BY-2cell lines are screened out. After treated with auxin2,4-D the green and cyan fluorescent enhanced apparently. It demonstrates that the auxin increase the activity of ABP1and CesA1. The vesicular transportation is also activated with a stronger inner fluorescent in the cell. It confirms our predication that auxin accelerates the vesicular transportation and transfer more cellulose synthase to the cell membrane. The more Cellulose Synthase in cell membrane can lead more cellulose synthesis in the wall thus the cell elongated.