Figure 1

Splicing of endogenous VEGF gene and design of GFP reporter minigene for assessing VEGF exon 6A splicing. A. Exon utilization in VEGF165 and VEGF189. Both VEGF165 and VEGF189 use exons 1 through 5 plus exon 7 and 8. The mRNA for VEGF189 also contains the alternatively spliced exon 6A. B. Design of the GFP reporter minigene for assessment of VEGF exon 6A usage. The minigene consists of a GFP gene driven by cytomegalovirus (CMV) promoter which is interrupted by a shortened version of VEGF intron 5, exon 6A and a shortened version of intron 6. Detailed description of the components is given in Methods. There are two possible splicing reactions for the transcripts of pGFP-E6A. If the splicing pattern is like that in VEGF189, exon 6A is included so as to interrupt the GFP reading frame. If the splicing pattern is similar to that in VEGF165, exon 6A is excluded and a complete GFP open reading frame is generated giving rise to active GFP protein. The arrows indicated the position of primers used for amplification of spliced mRNA products.