Fig. 4

Fig. 4 MO-ATG depletes full-length tnnt2, while MO-E13 generates truncated tnnt2. (A) Schematic illustration of the zebrafish tnnt2 gene. Exons are indicated by black boxes, with the alternatively spliced exons indicated by asterisks. The exons that encode an N-terminal Tm binding domain and a C-terminal Tm/tnnc/tnni binding domain are underlined (Watkins et al., 1995). The sites targeted by the MOs are indicated with arrows. For more detailed annotation, see Supplemental Table S1. (B) Quantification of cardiac contractility in WT and morphants. Shown are mean ± s.d. of the shortening fraction of ventricular chambers. In contrast to a silent heart in MO-ATG morphants, weak contractility still exists in MO-E13 morphants. For video, see the Supplemental Movies. *p < 0.01, if compared with WT. N, number of hearts quantified. (C) Shown are embryos of WT, morphants of MO-ATG or MO-E13 at 48 hpf after immunostaining to reveal tnnt2, Tm, and tnni, respectively. Both tnnt2 and tnni are severely reduced in MO-ATG morphants while Tm is weakly reduced. tnnt2 is weakly reduced in the MO-E13 morphant while Tm and Tnni remain unchanged. Arrows indicate the ventricular chambers. Scale bar = 20 μm. (D) RT-PCR analysis of tnnt2 transcripts in MO-E13 morphants at 48 hpf. Primer pair 1 reveals a shorter product in the MO-E13 morphant (indicated by a white arrow), indicating an exon-skipping event that links exon 12 to exon 14. Primer pair 2 targeting at exon 8 detected an increased level of tnnt2 mRNA in MO-E13 morphants. The specific PCR product that is amplified by primer pair 2 is indicated by a black arrow. A primer pair targeting 18 S rDNA was used to optimize the starting amount of PCR template.