Pulsed-field gel electrophoresis (PFGE) is a standard typing method for isolates from Salmonella outbreaks and epidemiological investigations. Eight hundred sixty-six Salmonella enterica isolates from eight serotypes, including Heidelberg (n = 323), Javiana (n = 200), Typhimurium (n = 163), Newport (n = 93), Enteritidis (n = 45), Dublin (n = 25), Pullorum (n = 9), and Choleraesuis (n = 8), were subjected to PFGE, and their profiles were analyzed by random forest classification and compared to conventional hierarchical cluster analysis to determine potential predictive relationships between PFGE banding patterns and particular serotypes. Cluster analysis displayed only the underlying similarities and relationships of the isolates from the eight serotypes. However, for serotype prediction of a nonserotyped Salmonella isolate from its PFGE pattern, random forest classification provided better accuracy than conventional cluster analysis. Discriminatory DNA band class markers were identified for distinguishing Salmonella serotype Heidelberg, Javiana, Typhimurium, and Newport isolates.