Figure 3.

Expression of M₂ mAChR mRNA during zebrafish embryonic development analysed by RT–PCR and Southern blot. DNase I treated total RNAs from the indicated developmental stages were used as a template for the RT–PCR reaction. Equal aliquots of cDNA were amplified with oligonucleotides specific for M₂ mAChR (582 b.p. product) or α-actin (341 b.p. product). Five μl of the RT–PCR reaction products were loaded on a 1.2% agarose gel and stained with ethidium bromide as shown in first panel (M₂) and second panel (α-actin). Southern blot analysis using a DIG-labelled M₂ mAChR DNA probe was carried out to confirm the identity of the amplified DNA as shown in the third panel. Hour postfertilization (h.p.f.) or days postfertilization (d.p.f.) of the embryo stage are shown on the upper edge.