RT-PCR analysis reveals the presence of introns in various cDNAs. (A) (Left panel) PCR was performed on the FPGS gene using 2 sets of forward and reverse primers. (i) Primers residing within the first and third exons (EX1-up and EX3-dw). RT-PCR on cDNA from CCRF-CEM cells produced the expected product size (eg, 263 bp, lane 1), whereas the MTX^R5P cells cDNA yielded a markedly longer product of 1570 bp (lane 2). (ii) PCR was performed to confirm the presence of intron 10 in cDNA from MTX^R5P cells using a forward primer (INT10) residing within this intron and EX13-dw (lane 4). The white vertical line has been inserted to indicate repositioned gel lanes. (Right panel) Scheme illustrating the positions of the primers as well as the predicted and observed PCR products. (B,C) PCR was performed on various genes using a panel of forward and reverse primers (Table 3) residing within neighbor exons to obtain different product sizes revealing normal and unspliced cDNAs. (B) RT-PCR on cDNA from parental and MTX^R5P cells produced the expected product sizes. (C) RT-PCR on WT cDNA produced normal size products, whereas MTX^R5P cell's cDNA revealed longer fragments as predicted for unspliced cDNA containing introns; all sizes of PCR fragments are depicted in Table 3.