Whole-mount in situ hybridization (WISH) was performed as previously described (Pearson et al., 2009; (link)
Sasidharan et al., 2013; (link)
King and Newmark, 2013 (link)). miRNA-based WISH was carried out using digoxigenin-labeled miRCURY LNA probes obtained from Exiqon. For immunostaining, animals were fixed using Carnoy's solution as described (Sánchez Alvarado and Newmark, 1999 (link)). Rabbit anti-ARRESTIN (1:5000, clone VC-1; gift of Dr Kiyokazu Agata, Kyoto University, Japan) and mouse anti-SYNORF-1 (1:100, DHSB) primary antibodies were used. Species-specific secondary antibodies were obtained from Molecular Probes (1:400). Hoechst 33342 (25 µg/ml; Sigma) was used as a nuclear counterstain. Animals were mounted with Mowiol mounting medium (Sigma) and stored at 4°C until imaging.
Sasidharan et al., 2013; (link)
King and Newmark, 2013 (link)). miRNA-based WISH was carried out using digoxigenin-labeled miRCURY LNA probes obtained from Exiqon. For immunostaining, animals were fixed using Carnoy's solution as described (Sánchez Alvarado and Newmark, 1999 (link)). Rabbit anti-ARRESTIN (1:5000, clone VC-1; gift of Dr Kiyokazu Agata, Kyoto University, Japan) and mouse anti-SYNORF-1 (1:100, DHSB) primary antibodies were used. Species-specific secondary antibodies were obtained from Molecular Probes (1:400). Hoechst 33342 (25 µg/ml; Sigma) was used as a nuclear counterstain. Animals were mounted with Mowiol mounting medium (Sigma) and stored at 4°C until imaging.
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