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Figure 2. Expression of UFS genes and immunolocalisation of UFS proteins. (A) RT- PCR analysis of a developmental stage series derived from whole embryo RNA. hpse2 was absent at Nieuwkoop-Faber (NF) stage 1 but was detected from stage 15 to stage 45. lrig2 was expressed at stage 1, then weakly at stages 15/20, after which transcript levels became prominent. hpse1 was detected at all stages analysed. All three transcripts were present in adult urinary bladder (Blad). gapdh was included as a cDNA loading control. In the H2O column, no cDNA was used. (B) Transverse section through the trunk of a stage 40 embryo, with the dorsal surface uppermost. Brown colour shows positive IHC signal for heparanase 2, most prominent in the myotomes flanking the central neural tube, and fainter expression in the ventrolateral neural tube. (C) Adjacent section to (B), with the same orientation, showing minimal IHC signal after preincubating primary antibody with the immunising peptide. (D) Transverse section through the trunk at stage 30, with the dorsal surface uppermost. Prominent heparanase 2 immunoreactivity in detected in the ventrolateral regions of the neural tube (NT), in the notochord (Nc) and in the flanking skeletal muscle myotomes (My). (E) Adjacent section to (D), with the same orientation, immunostained for LRIG2. A similar pattern to that of heparanase 2 is observed, extending into the dorsal half of the neural tube. (F) Transverse section from a stage 42 embryo, with the left side uppermost. Three sections of gut are apparent, the archenteron having undergone coiling. Heparanase 2 immunoreactivity is observed in the luminal surface of the gut epithelium, and shown in closer detail in (G). B-G were counterstained with haematoxylin, rendering nuclei blue. Scale bars are 50 μm. |
