Genetic investigation and phylogenetic analyses of 21 species, representing 8 genera of the subfamily Malvoideae, from Saudi Arabia were carried out by using Start Codon Targeted (SCoT), the Inter Simple Sequence Repeats DNA (ISSR), and united ISSR and SCoT markers. Ten SCoT and five ISSR primers created 149 polymorphic enlarged fragments, which jagged to a comparatively high level of genetic difference in Malvoideae. SCoT markers exposed a higher level of polymorphism (89 bands) than ISSR (60 bands). Gathering of genotypes within groups was greatly similar when SCoT and ISSR consequent dendrograms were compared. Five clusters and clades can be documented within Malvoideae, which regularly verify, but also partially deny, traditional groupings. Taxonomic and phylogenetic consequences are debated in contrast with the existing morphological and phylogenetic data. The results of this study offer useful data for evaluating the taxonomy of Malvoideae at both tribes, infrageneric, and subgeneric levels. In general, the results reported here are largely consistent with the phylogenetic findings that Abutilon, Hibiscus, and Malva are polyphyletic. Also, species of sections Bombicella and Malva are highly heterogeneous. The most exciting result of this analysis was identifying Senra incana with unique characters and telling that it should be preserved as a separate tribe. In the similar way, distinctive variances between the closely related genera Fioria and Althaea were also noted suggesting that they should be placed in different tribes. Furthermore, the present results indicate some level of parallel among the species of Pavonia and support the monophyly of this genus.