A conventional source of natural rubber (NR) is from Hevea (Hevea brasiliensis). NR production from Hevea is limited by its narrow growth region and susceptibility to leaf blight disease. In response to these limitations, and to address NR supply and security, guayule (Parthenium argentatum Gray) has been developed as an alternative crop. NR consists mainly of cis-1,4-polyisoprene synthesized in/on rubber particles (RPs). RPs resemble lipid droplets (LDs) in that both are generated from endoplasmic reticulum (ER) and consist of hydrophobic cores and phospholipid monolayers. A prominent ER-related protein seipin plays an essential role in controlling LD biogenesis from the ER surface. Prior expression of AtSEIPIN1 in Arabidopsis increased LD size and consequently increased its core constituent triacylglycerol content in seed. To investigate if AtSEIPIN1 can similarly impact RPs and production of cis-1,4-polyisoprene, we generated three independent transgenic guayule lines overexpressing AtSEIPIN1 (SEIoe). Compared with wild-type, fewer RPs were observed, and RP size was significantly reduced in all SEIoe lines. Although not always statistically significant, NR quantity and quality were also reduced in SEIoe lines. Resin contents were reduced in mature stems, but not in young plantlets of SEIoe lines. The mechanisms of how AtSEIPIN1 influences RP biogenesis and NR production in guayule are discussed.