Figure 3.

Validation of novel PROM1 TSS using 5′ RACE PCR. (A) Schema illustrating primer design for PROM1 TSS validation. PROM1 TSS is amplified using a pair of outer and inner primer sets targeted specifically to 5′ Rapid amplification of cDNA ends (RACE) adapter and PROM1 sequences. (B) The 5′ end regions of PROM1 mRNA were amplified from CD133⁺ and CD133⁻ populations of LM-Mel-34 cell lines. Gel electrophoresis of amplified products shows difference in expression level in CD133⁺ and CD133⁻ populations. Amplified 5′ends of PROM1 RNA were cloned in to plasmid pcDNA3.1 and then sequenced. (C) UCSC genome browser view of sequenced 5′ end region of PROM1 mapped to the promoter. The tracks shown here illustrates CAGE captured TSS regions from CD133⁺ melanoma cell lines and the RACE PCR identified 5′ end of the PROM1 transcript. The other tracks show outer and inner RACE PCR primers specific to PROM1.