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ZFIN ID: ZDB-FIG-190218-15
Yan et al., 2018 - Maternal Huluwa dictates the embryonic body axis through β-catenin in vertebrates. Science (New York, N.Y.)   362(6417) Full text @ Science
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Anatomical Terms:
Stage Range: 2-cell to Adult
PHENOTYPE:
Fish:
Observed In:
Stage Range: 4-cell to Adult

Fig. 2 Identification and verification of tsu01sm mutant gene. (A) Top: Illustration of the zebrafish hwa wild-type and tsu01sm mutant alleles. Arrowheads indicate the positions of PCR primers. The green arrow indicates the translation start site. Bottom: Electrophoretic result of PCR products in wild-type, heterozygous, and homozygous mutant embryos. M, molecular weight markers. (B) hwa mRNA detection by WISH (left) and by qRT-PCR (right). bb, Balbiani body in stage I oocyte. For qRT-PCR, oocytes are a pool of stage I–III oocytes and eggs are squeezed from females. The relative hwa mRNA levels are averages (±SEM) from three independent experiments, normalized to eif4g2a levels. ***P < 0.001. Scale bars, 100 μm. (C) Mhwatsu01sm mutant embryos form the body axis or are dorsalized by hwa overexpression. Left: Morphology of classified embryos at 24 hpf. Right: Ratios of embryos in different classes. (D) Induction of secondary body axis by ectopic hwa mRNA overexpression. One blastomere of 32-cell stage Tg(gsc:EGFP) transgenic embryos or two opposite blastomeres of 32-cell stage Tg(gsc:EGFP);Mhwatsu01sm mutant embryos were injected with 50 pg of hwa and 50 pg of mCherry mRNAs, as illustrated at left. Embryos were observed dorsally (top) or laterally (bottom) at 6 hpf under a fluorescence dissection microscope, and those with two dorsal organizers (EGFP-positive) were observed again at 24 hpf. The ratio of embryos with double organizers is indicated. Scale bars, 100 μm. (E) Generation of a hwa mutant allele by Cas9 knockout. Top: Illustration of the mutant allele with an underlined premature stop codon. Bottom left: A group of Mhwatsu32c mutant embryos at 24 hpf. Bottom right: Alteration of the dorsal markers gsc and chd and the ventral marker eve1 at shield stage; scale bar, 100 μm.

Gene Expression Details
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay
chrd WT standard conditions Shield shield ISH
hwatsu32c standard conditions Shield Not Detected shield ISH
eve1 WT standard conditions Shield whole organism ventral region ISH
hwatsu32c standard conditions Shield whole organism ventral region ISH
gsc WT standard conditions Shield shield ISH
hwatsu32c standard conditions Shield Not Detected shield ISH
hwa WT standard conditions 4-cell whole organism ISH
Adult oocyte stage I mitochondrial cloud ISH
hwatsu01sm standard conditions 2-cell whole organism RTPCR
4-cell Not Detected whole organism ISH
Adult oocyte RTPCR
Adult Not Detected oocyte stage I mitochondrial cloud ISH
Adult oocyte stage V RTPCR
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
hwatsu01sm standard conditions 4-cell whole organism hwa expression absent, abnormal
2-cell whole organism hwa expression decreased amount, abnormal
Prim-5 whole organism morphology, abnormal
Prim-5 yolk split, abnormal
Adult oocyte hwa expression decreased amount, abnormal
Adult oocyte stage I mitochondrial cloud hwa expression absent, abnormal
Adult oocyte stage V hwa expression decreased amount, abnormal
hwatsu32c standard conditions Shield shield chrd expression absent, abnormal
Shield shield gsc expression absent, abnormal
Shield whole organism ventral region eve1 expression increased distribution, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Science (New York, N.Y.) for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Science