The sequences and maps of the Chlamydomonas artificial miRNA vectors pChlamiRNA2, pChlamyRNA3 and pChlamyRNA3int
- pGR106 (Vector NTI)
- pGR106 (Sequence, Word format)
- pGR106 diagram (Powerpoint format)
- pGR106 polylinker seq
- pGR107 polylinker seq
Suppressors of Gene Silencing
The 35Spro-35Ster expression cassette of pJIT61 was excised as a kpnI-XhoI fragment and mobilised into the pBin19 binary vector T-DNA. The cassette was inserted into KpnI-SalI digested pBin19, therefore eliminating the SalI site. This led to plasmid pBin61. You will find the sequence of the vector as a MS Word file below.
Inserting suppressors of gene silencing into pBin61:
The SmaI site in pBin61 was used to clone all the viral suppressors in sense orientation. Blunt-end PCR-amplified fragments of the suppressors were used for cloning. It is therefore not possible to excise the suppressor sequence from the pBin61 vector. You will find sequences of each individual suppressor constructs cloned in pBin61 below.
Rx Resistance Gene
The two amplicon constructs are PUC19-based vectors. These vectors are as described in Cell, 2000 (103), 157-167. PVX-GFP-*CP has a deletion spanning the entire coat protein. PVX-GFP-*TGB-*CP is based on PVX-GFP-*CP in which the entire trible-gene-block encoding the movement proteins (including the p25 suppressor of PTGS) has been removed. Both amplicon constructs are under 35S promoter and Nos terminator. The expression cassettes can be mobilised with a SacI digest and easily subcloned in any binary vector.