Zebrafish DND1 (drDND1) counteracts inhibition of TDRD7 by miR-430 through binding to adjacent URRs. (A) One-cell-stage zebrafish embryos were injected with dead end morpholino or control morpholino. RNA was extracted and subjected to Q- RTPCR analysis to compare endogenous levels of TDRD7 and Vasa to Odc, and TDRD7 to Vasa. (B) One-cell-stage zebrafish embryos were co-injected with DsRed-TDRD7-3′UTR (3′TDRD7) and gfp-vasa-3′UTR (3′vasa) together with dead end morpholino or control morpholino. (C) One-cell-stage zebrafish embryos were co-injected with RNA containing the yfp open reading frame fused to the wild-type TDRD7 3′ UTR (3′TDRD7wt), RNA containing the cfp open reading frame fused to the miR-430 binding site mutated TDRD7 3′ UTR (cfp- 3′TDRD7mut1) and vasa-dsRed (for labelling the germinal granule for easier identification of germ cells) together with dead end morpholino or control morpholino. (D) RNA containing the DsRed open reading frame fused to the wild-type nanos-1 3′ UTR (3′nos1wt) was co-injected into one-cell-stage zebrafish embryos together with RNA containing the gfp open reading frame fused to different versions of the TDRD7-1 3′ UTR. The different TDRD7 3′ UTR that were used were: wild-type TDRD7 3′UTR (3′TDRD7wt) or TDRD7 3′UTR where the putative DND1 interaction sequence was mutated (3′TDRD7mut2). Error bars depict the standard error of the mean (SEM), p-value was calculated using Ttest.
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