Validation of DNMT3A splice variant expression. Validation of DNMT3A splice variant expression. (A) KD and (B) OE of DNMT3A transcripts (Tr.) by RT-qPCR relative to control samples scrambled and empty, respectively (KD and OE each in independent biological replicates, n = 3; mean ± SD). Significance was estimated by the t test; *P < 0.05, **P < 0.01, ***P < 0.001. (C) Western blot analysis of DNMT3A isoforms upon OE. Please note that the N-terminal antibody targets only transcripts 1+3 and 4 (expected at ∼130 kD and 18 kD, respectively). Actin was used as a reference protein. The Western blot was performed with limited cell numbers, and protein levels, therefore, differ between samples. Protein validation upon OE of Tr.2 and KD of all transcripts was not possible because of limited cell amount and unreliable antibody targeting the C-terminus (results not demonstrated). Tanja Božić et al. LSA 2018;1:e201800153 © 2018 Božić et al.