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Fig. S2 Effects of cdt1 morpholino on morphology and mRNA splicing. (A) Brightfield microscopy images demonstrate the morphology of 24-hpf zebrafish embryos after injection of the indicated morpholinos. Note that the knockdown of p53 alleviates small head and shorter body axis phenotype in cdt1 and/or emi1 morphants. (B) Brightfield microscopy images demonstrate the 24-hpf morphological phenotypes due to injecting cdt1 MO into embryos wildtype or mutant for emi1. The quantitation on the right illustrates lack of morphological rescue of emi1 defects by cdt1 morpholino. (C) Cdt1 morpholino injection results in aberrant splicing of cdt1 transcripts (mainly partial insertion of intron 2). RT-PCR analysis was performed with RNA from pools of 20 embryos injected with the indicated morpholinos. Splicing of cdt1 was analyzed using primers in exon sequences that surround the target exon 3 (top panel). Inclusion of intron 3 in transcripts was assayed using primers targeting exon 3 (forward) and intron 3 (reverse) sequences. Note the (exon 3– intron 3) background product in control MO-injected embryos, which could results from unspliced transcript or genomic DNA contamination. Knockdown of cdt1 did not affect the emi1 splicing defects caused by emi1 morpholino (third panel form top). Co-injection of p53 morpholino did not alter the splicing patterns of either cdt1 or emi1 transcripts. RT-PCR of beta (β)-actin was used as a control for RNA quality and quantity (bottom panel).