Species-specific RPL13A region sno-lncRNA is derived from species-specific alternatively spliced rpl13a transcripts. (A) De novo transcript assembly revealed previously uncharacterized alternative spliced rpl13a transcripts (mouseR1_pAplus_cufflinks). At least two new rpl13a isoforms (back arrows on right) were identified to splice out an intron containing SNORD33 and SNORD34. Y-axis, normalized read densities of poly(A)-/ribo- RNA-seq (red) and poly(A)+ RNA-seq (black) of mouse ESC and hippocampus transcriptomes. (B) No such alternative spliced rpl13a transcripts in human ESC H9 and HeLa cells from both de novo assembly (h9_pAplus_cufflinks) and known annotation (blue lines). Y-axis, normalized read densities of poly(A)-/ribo- RNA-seq (red) and poly(A)+ RNA-seq (black) of human H9 and HeLa transcriptomes. (C) Validation of alternative spliced rpl13a transcripts by RT-PCR. Left, the previously uncharacterized alternative splicing event of RPL13A gene could only be detected in mouse, but not in human, with RT-PCR. Right, a schematic drawing of different splicing events and their validation primers in RPL13A. Black bars, RPL13A exons; Blue circles, snoRNAs; Red arrows, PCR primer sets. (D) The alternative splicing of RPL13A causes amino acid changes at the C-terminal of RPL13A protein. Top, schematic diagram of canonical splicing of RPL13A and its amino acid sequence; Bottom, schematic diagram of uncharacterized splicing of RPL13A and its amino acid sequence.