| 質粒簡介 |
Tobacco rattle virus (TRV) is a bipartite, positive-sense RNA virus. TRV1 (i.e., RNA1) encodes 134 and 194 kDa replicase proteins from the genomic RNA, 29 kDa movement protein, and 16 kDa cysteine-rich protein from subgenomic RNAs. TRV1 can replicate and move systemically without TRV2 (i.e., RNA2). TRV2 encodes coat protein from the genomic and two nonstructural proteins from subgenomic RNAs (A). To develop the TRV-based VIGS vector, a cDNA clone of TRV1 or TRV2 (Ppk20 strain) was placed between a duplicated CaMV 35S promoter (2X35S) and nopaline synthase terminator (NOSt) in an Agrobacterium T-DNA vector. In the TRV2 cDNA construct, the nonessential structural genes were replaced with MCS for cloning the target gene sequences. Further, a self-cleaving ribozyme site was included at the 3' end of TRV2. The TRV1 (AF406990)-based viral vector, pTRV1 (linear plasmid 6.791 kb), is used along with pTRV2 for silencing. The pTRV2 has three variants. The first one is suitable for conventional cloning (AF406991, linear plasmid 9663 nucleotide). The second variant is a gateway-cloning-compatible vector. The third variant is suitable for ligation-independent cloning. Rz, self-cleaving ribozyme; MCS, multiple cloning site; CP, coat protein; MP, movement protein. These vectors were created by Dr. S.P. Dinesh-Kumar, UC Davis, USA. Conventional cloning-compatible (YL156) and ligation-independent-compatible (YY13TRV2) vector plasmid DNA can be obtained from ABRC, Stock No. CD3-1040 vector name YL156. Ligation-independent-compatible TRV2 vector plasmid DNA is ABRC Stock No. CD3-1042, vector name YY13. |
