Technical Abstract: Several proteins have been identified and implicated in natural rubber biosynthesis, one of which, the small rubber particle protein (SRPP), was originally identified in Hevea brasiliensis as an abundant protein associated with cytosolic vesicles known as rubber particles. While previous in vitro studies suggest that SRPP plays a role in rubber biosynthesis, in vivo evidence is lacking to support this hypothesis. To address this issue, a transgene approach was taken in Taraxacum kok-saghyz (Russian dandelion or Tk) to determine if altered SRPP levels would influence rubber biosynthesis. Three dandelion SRPPs were found to be highly expressed on dandelion rubber particles. The most abundant particle associated SRPP, TkSRPP3, showed temporal and spatial patterns of expression consistent with patterns of rubber accumulation in dandelion. To confirm its role in rubber biosynthesis, TkSRPP3 expression was altered in Russian dandelion using over-expression and RNAi methods. While TkSRPP3 over-expressing lines had slightly higher levels of rubber in their roots, relative to controls, TkSRPP3 RNAi lines showed significant decreases in root rubber content and produced dramatically lower molecular weight rubber than control lines. Not only do results here provide in vivo evidence of SRPP affecting the amount of rubber in dandelion root, but they also suggest a function in regulating the molecular weight of the cis-1,4-polyisoprene polymer.