3 race kit

3’-RACE PCR was carried out using the 3’-RACE Kit (Takara D314) following the manufacturer's instructions. The PCR relied on the nested adaptor primer and specific primers for the first exon of LOC_Os06g44034. Relevant PCR primer sequences are given in S4 Table.

Total RNAs were extracted from the human prostate tumors and the normal tissues and various cell lines using Trizol (Life Technologies). Reverse transcription was performed using the M-MLV reverse system (Takara) to obtain cDNA. qPCR was performed using with the SYBR Premix Ex Taq mix (TaKaRa) according to the manufacturer’s instructions, and the samples were run on an iQ5 Multicolor Real-time PCR Detection System (Bio-Rad). Results were normalized to expression levels for reference genes (GAPDH or EIF4H). The relative expression levels of genes were calculated using the ∆∆CT method. 3′RACE kit (Takara) was used to convert RNAs of the prostate tumors into cDNAs by a reverse transcriptase and oligo-dT adapter primer. The cDNAs were amplified by using FOXP2-specific primers, which annealed to exon 6, exon 11, and exon 16, and an oligo-dT adapter primer, respectively. Amplified fragments of the putative sizes were subjected to Sanger sequencing. Genomic DNAs of the fusion-positive tumors (PC_1 and PC_11) were used for long-range PCR. Amplified fragments of the putative sizes were subjected to Sanger sequencing. The primers for RT-PCR, qPCR, 3′RACE-PCR, and long-range PCR are available in Supplementary file 1f.

A SMARTer^® RACE 5′/3′ Kit (Clontech) was used to determine the 5′ end of AS1eRNA. The 3′ end of AS1eRNA was determined by using a 3′ RACE kit (TaKaRa). All RACE primers are listed in Supplementary Table S5.