Unexpected variations can occur during natural and in vitro propagation of bananas (banana and plantain) to generate off-types. The molecular basis of such variations is not well-understood. This study aimed to characterize the functions of genomic regions varying within clones grown naturally or in vitro. Fifty-four simple sequence repeat (SSR) markers and six primer combinations of EcoR I/Msp I-amplified fragment length polymorphism (AFLP) were used to analyze accessions of the AA, BB, AB, AAA, AAB, and ABB groups of Musa, and polymorphic regions were sequenced to characterize candidate genes. One SSR locus with significant similarity to an arcelin gene revealed a deletion in a subculture regenerant. In AFLP analysis, 24 (6.15%) of 390 bands accounted for within-clone variations, with 0.5% and 5.65% occurring in natural and in vitro propagated plants, respectively. Sequence homology searches revealed that most polymorphic regions were related to cytochrome P450, cell-wall biosynthesis, and senescence genes. The importance of these candidate genes is discussed. The plants harboring the variations were field-established to relate molecular variations to phenotypic changes. Sixteen of the sequences registered in Genbank (ET165586 to ET165601) and select PCR primers from this study can be further tested for variations between normal clones and off-types in Musa.