Figure 5

RT-PCR analysis of potential mesothelin exon 16–17 splicing differences between transcript variants 1 and 3. A. Diagram of the PCR strategy, showing predicted size of amplified products from transcript variants 1 and 3. PCR primers (Table 1, line 3) are indicated by arrows. B. Agarose gel analysis of representative RT-PCR results. Lane M, molecular size markers; Lane 1, no template PCR control; Lane 2, PANC-1; Lane 3, AsPc-1; Lane 4, HeLa; Lane 5, OVCAR-3; Lane 5, normal ovary tissue sample; Lane 6–8, three individual primary ovarian tumors. The product with a size between 262 and 344 bp represents heteroduplex formation between transcript variants 1 and 3 (tr1, tr3). C. RT-PCR and agarose gel analysis of full length mesothelin (transcript 1 versus 3) in cytoplasmic (C) versus nuclear (N) RNA fractions as a template, using primers in Table 1, line 4. Lane M, molecular size marker; Lane 1, OVCAR-3 cytoplasmic fraction; Lane 2, OVCAR-3 nuclear fraction; Lane 3, AsPC-1 cytoplasmic fraction; Lane 4, AsPC-1 nuclear fraction; Lane 5, HeLa cytoplasmic fraction; Lane 6, HeLa nuclear fraction; (--), no template PCR control.