XB-IMG-86835
Xenbase Image ID: 86835
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Figure 4. Morpholino knockdown of heparanase 2. (A) Schematic diagram of the Xenopus tropicalis hpse2 gene, showing: exons (blue blocks); splice MO targets (red blocks) at the splice acceptor (MO1) and splice donor (MO2) sites of exon 2; PCR primers flanking exon 2 (black arrows); and the premature stop codon in exon 3 (red asterisk) generated by MO1. (B) Injection of increasing amounts of MO1 induced mis-splicing of hpse2, with 5 ng abolishing expression of wild type (wt) hpse2 mRNA in favour of a shorter mRNA (exon 2δ). (C) Sanger sequencing of the shorter PCR product confirmed absence of exon 2 (indicated by dotted vertical line). A novel, in-frame premature stop codon was generated in exon 3 (asterisk). (D) Injection of 5 ng MO1 led to near complete loss of heparanase 2 neural tube and myotome immunoreactivity, as demonstrated in this stage 40 embryo; CTL MO on left and MO1 on right. (E) Frequency (%) of the hypomotility, lack of gut looping (gut defect) and tail curvature (tail defect) phenotypes associated with administration of CTL MO or MO1, with total numbers of embryos assessed indicated by ‘n’. (F) The upper two images depict CTL MO-administered embryos and the lower two images show effects of MO1. Left- hand section depicts embryos viewed from the side; note the protruding proctodeum (white arrow) and tail curvature in the morphant. The two frames on the right depict the embryos viewed from their ventral aspects; note that gut coiling is present in the control embryo but not in the morphant. Image published in: Roberts NA et al. (2014) © The Author 2014. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license
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