The effects of transcription initiation (ti)RNA sponges and mimics on p21 (cyclin-dependent kinase inhibitor 1A gene, also known as CDKN1A ) expression. (a) Transfection by the CCCTC-binding factor (CTCF) proximal (cp)-tiRNA sponge resulted in an increase in p21 mRNA and (b) p21alt transcript levels. Reverse transcription was non-specifically primed and cDNA was analyzed by quantitative (qPCR) using primer set 2 for p21 mRNA and primer set 1 for p21alt RNA. (c) Further validation of the results in (a) and (b) were confirmed by transfection with the cp-tiRNA sponge followed by strand-specific quantitative reverse transcription (qRT)-PCR which showed an increase in both antisense and sense p21alt transcript levels, and (d) an increase in sense p21 mRNA. (e) Transfection of cp-tiRNA mimics resulted in a decrease in both sense classical and alternate p21 transcripts. (a-e) Samples were analyzed at 72 h post transfection. The averages of samples transfected in triplicate are shown, error bars represent the standard errors of the means, and P values from paired t tests are shown.