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Table 2 Overview of the aphidicolin, actinomycin D and puromycin treatment experiments on Mnemiopsis leidyi embryos

From: A cleavage clock regulates features of lineage-specific differentiation in the development of a basal branching metazoan, the ctenophore Mnemiopsis leidyi

Developmental stage

Treatment

Observation

Conclusion

2-cell stage

Arrest of DNA synthesis for 2 h with aphidicolin, 2 μg/ml

No combs formed (0/30)

DNA synthesis is essential for comb formation.

16-cell stage

32-cell stage

60-cell stage

27/30 (90%) formed combs, comb formation was delayed by 2 h

4 hpf

29/30 (97%) formed combs, comb formation was delayed by 2 h

5 hpf

29/30 (97%) formed combs, comb formation was delayed by 2 h

60-cell stage and 4 hpf

Arrest of DNA synthesis and cytokinesis with aphidicolin, 2 µg/ml, plus cytochalasin B, 1 to 2 μg/ml

Immediately arrested, single nuclei observed, neither combs nor photoprotein were formed in any embryo (0/30)

DNA synthesis is essential for comb formation even in cleavage-arrested embryos.

1- to 2-cell stage

Blocking of transcription with actinomycin D, 10 μg/ml

0/30 gastrulated, development arrests at 32- to 60-cell stage

Zygotic transcription begins around the 32- to 60-cell stage and the specification of the combs requires more than only maternal RNAs.

4- to 8-cell stage

0/30 gastrulated, development arrests at 32- to 60-cell stage

16- to 32-cell stage

10/30 (33%) gastrulate, 20/30 (67%) arrest at 32- to 60-cell stage

60-cell stage

28/30 (93%) gastrulate but no pharynx is formed, 2/30 (7%) do not gastrulate

5 hpf (post-gastrulation)

No pharynx is formed

2-cell stage

Arrest of translation with puromycin, 125 μg/ml, 25 μg/ml

0/30 formed combs

Comb protein is translated prior to 6 hpf, comb formation requires protein that is translated in the embryo and not maternally loaded.

16-cell stage

0/30 formed combs

60-cell stage

0/30 formed combs

6 hpf (post-gastrulation)

32/30 (97%) formed combs