Fig 1. Validation of RP11-326A19.4/TCONS forms in HEK293T.

A, Manhattan plot of the 15q.26 CAD risk locus. Top, local Manhattan plot Data was drawn with LocusZoom using CAD association data from Nikpay et al [34]. Bottom corresponding genes and transcripts from GENECODE (green and blue) and Human Body Map (brown), visualized via the UCSC browser data, to scale with the Manhattan plot. B, splice form analysis of RP11-326A19.4. RNA was isolated from HEK293T and subjected to qRT-PCR analyses with primer sets targeted against various exons of the predicted transcripts. Top, schematic of primers used for PCR and alternating exon organization. Black bars represent exons shared by both spliced forms while orange bars represent additional exonic sequences unique to the TCONS_00023493 form. Size of predicted introns are indicated on top. Bottom, agarose gel of products. Primer pairs used are indicated above the gel. Predicted sizes of PCR fragments (in bp) are indicated under the gel and blue arrows point to the predicted migration position. In addition primers targeting exon 1 and 2 resulted in the presence of an additional band, longer by about 100 bp, consistent with the presence of an an alternative exonic sequence, situated between predicted exon 1 and exon 2.