1) Addition of an expression cassette with the miR173 precursor placed behind the UBQ11 constitutive promoter;
2) No promoter for expression of the MIGS construct. Instead, a promoter of choice can be cloned using the MCS just upstream of the attR1 site
Spectinomycin and Chloramphenicol
1) Before recombination Chloramphenicol might be used for bacteria selection (resistance lost after recombination). 2) E. coli DB3.1 strain (or some other ccdB survival strain) should be used for the amplification of the plasmid due to the presence of the lethal ccdB gene. After recombination, any common strain, such as DH5alpha is recommended. 3) Due to pGreen backbone characteristics, the helper plasmid pSOUP is required for replication of MIGS vector in Agrobacterium. PSOUP can be requested directly at http://www.pgreen.ac.uk/.
Please acknowledge the principal investigator and cite this article if you use
this plasmid in a publication. Also, please include the text "Addgene plasmid
35252" in your Materials and Methods section.