We have established a clonal porcine subcutaneous preadipocyte cell line for the first time. This cell line, named PSPA, shows a fibroblast-like phenotype and keeps on growing under growth medium even after reaching confluence. Adipose conversion occurs only when confluent cultures are stimulated with insulin, dexamethasone, biotin, pantothenate, and octanoate. Following exposure to this differentiation medium, growth was arrested immediately without mitotic expansion and PSPA cells exhibited a marked increase in intracellular lipid accumulation accompanied by changes in gene expressions directly related to fat metabolism and also in extracellular environments. Since PSPA cells were established, it has become possible to end the discussion over whether there is species specificity in the adipose conversion between pig and mouse by comparing PSPA cells and 3T3-L1 cells, the most widely studied preadipocyte cell line. Consequently, we demonstrated that responsiveness to inducers and fatty acid composition was unequal, despite being treated with the same media. Thus, species-specific differences indeed exist in adipogenesis between these two, meaning that PSPA cells would be an in vitro model specialized to improve understanding of pig adipocyte growth and differentiation in controlling the carcass fat content of pigs.