Abstract
Gateway system is efficient and rapid cloning technique to transfer the target DNA fragments into vectors. It seems to be easy that cDNA library made by Gateway system is able to apply to library construction based on various kinds of vectors. First, we constructed Arabidopsis cDNA library utilizable for Gateway system. The double-stranded cDNA with attB1 and attB2 sites was synthesized from Arabidopsis green leaves, normalized by DSN treatment, and then cloned into pDONR vector. Using this normalized cDNA library, we made the library based on VIGS vector derived from TRV by Gateway system. Furthermore, we have tried to establish the phenotype-dependent rapid screening system using this VIGS library. Each clone of Agrobacterium harboring VIGS library was infiltrated into leaves of 3-weeks-old Arabidopsis, and then the resulted phenotype were observed. We'll report the detail results of screening for cell death-associated genes and of application toward screening based on various phenotypes.
