Figure 5. Minigene constructs and products obtained in the in vitro splicing assay. A: Shown is a schematic representation of the constructs, which include PROM1 exons 21, 22, and 23 (represented by boxes), flanked by intronic sequences (represented by straight lines). The deletion breakpoint is indicated by a dashed line in the mutant constructs (del/C and del/G), while the deleted bases are indicated by a dashed box in the corresponding wild-type (wt) constructs (+/C and +/G). Either a C or a G is present at position −6 of intron 21. Also shown are the locations of primers used for RT–PCR analysis (indicated by arrows). B: Constructs were transfected into COS-7 cells, followed by RNA extraction and RT–PCR analysis. cDNA derived from wt constructs yielded two products, with and without exon 22. cDNA derived from constructs harboring the deletion yielded only the ex22- product.