Figure 3
Expression of naPINK1, PINK1 and svPINK1 following knockdown of naPINK1 with short interfering RNA (siRNA) (a-d). Total RNA was isolated from neuroblastoma cell lines SK-N-MC and SH-SY5Y treated with two different siRNAs towards naPINK1 or an siRNA control which avoids targeting any known human sequence. Random hexamers were used in cDNA-synthesis when not otherwise stated and gene expression was determined using qRT-PCR. Data are presented as mean ± SE of the percentage mRNA abundance related to control siRNA treated samples in each experiment. (a) naPINK1 expression following AS-siRNA knockdown (n = 12 for control siRNA and AS siRNA 1, n = 11 for AS siRNA 2, one sample was excluded due to failed transfection). (b) PINK1 expression following AS-siRNA knockdown (n = 12 for control siRNA and n = 23 for pooled results from the two siRNAs). (c) svPINK1 following AS-siRNA knockdown (n = 12 for control siRNA and AS siRNA 1, n = 11 for AS siRNA 2). (d) To assess if naPINK1 and svPINK1 were polyadenylated and to exclude amplification of pre-mRNA species, naPINK1 and svPINK1 expression was determined in a subset of samples (n = 6) from which an additional cDNA synthesis was performed, this time using oligo d(T)16. Absolute values obtained for gene expression were similar to the random hexamer protocol while the extent of the interaction between naPINK1 and svPINK1 was arguably clearer.