Extended Data Fig. 3. Analyses of TBXT isoforms.

a, In vitro differentiation of human and mouse ESCs for inducing TBXT/Tbxt expression. Human and mouse ESCs differentiation assay was adapted from Xi et al. (2017)³⁶ and Pour et al. (2019)³⁷, respectively. Schematic adapted from icons created by Marcel Tisch via bioicons.com. b, Quantitative RT-PCR (RT-qPCR) of TBXT and MIXL1 expression during hESC differentiation, indicating correct induction of mesodermal gene expression program³⁶. c, Quantitative RT-PCR of Tbxt expression during mESC differentiation. Data in b and c were presented as mean +/− standard deviation of the relative gene expression levels. Sample number n = 3 represents RT-qPCR results from three biologically independent RNA samples, with each data point averaged from 3 technical replicates in quantitative PCR. d, RT-PCR of TBXT/Tbxt transcripts in human and mouse differentiated ESCs, highlighting the Δexon6 splicing isoform unique to human. RT-PCR results were presented as biological duplicates. e, Protein sequence alignment of TBXT-exon6 region in the representative mammals. All presented animals have tails except human and chimpanzee. f, The exon 6-coded peptide of Tbxt protein overlaps with large fractions of two transcription regulation domains. TA, transcription activation; TR, transcription repression. Functional domain annotation of mouse Tbxt protein was adapted from Kispert et al. (1995)⁹.