Figure 3. Changes in alternative splicing of RP11–295G20.2 were observed in Pnn-knockdown cells, and RT–PCR and in situ hybridization confirmed in addition to tRP11–295G20.2 expression in the human corneal epithelia. A: RP11–295G20.2 contains six exons spanning just greater than 6 kb. Corneal epithelial cells express RP11–295G20.2 isoforms containing exons 1, 4, 5, and 6. See Appendix 3. B: Knockdown of Pnn resulted in relative decreases in the expression of the two predominant isoforms containing exons 1–6 or 1, 4, and 6, and increased levels of isoforms containing 1, 4, 5, and 6. C: Real-time PCR (RT–PCR) of RNA from explant cultures from central corneal epithelia and limbal epithelia demonstrated robust expression of RP11–295G20.2. D: In situ hybridization analyses on human cornea sections revealed that a lncRNA RP11–295G20.2 is expressed in corneal and limbal epithelium with a nucleus-restricted pattern.