The present invention includes a method for detecting and isolating
sugarcane proteins that interact with the HC-Pro and P1 proteins of SrMV
and other proteins involved in gene silencing, particularly in sugarcane.
The method uses a two hybrid assay with an HC-Pro, P1, or other
silencing-related protein-containing bait protein and a prey protein
containing a polypeptide encoded by a DNA molecule in a cDNA library. The
method also includes identification of false positives through reverse
two-hybrid assays and using in vitro techniques such as farwestern blots
or pull down assays where plant physiological conditions may be
replicated. Finally, interactions may be confirmed in planta. Some novel
proteins used in and discovered using the these methods are also
identified. Methods of using viral and plant proteins to regulate
silencing in plants such as sugarcane are also discussed.