31 research outputs found

    Genomic target sequences for generating <i>tsp-15</i> knockin and <i>csnk-1</i> knockout strains using the CRISPR/Cas9 method.

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    ND: not determined. For tsp-15 repair template, the letter in red was for introducing the missense mutation and letters in blue were for introducing silent mutations. (TIFF)</p

    <i>csnk-1(lf)</i> and <i>skn-1(gf)</i> mutations exhibit synergy on the Sisi phenotype in 50 mM NaI.

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    csnk-1(lf) and skn-1(gf) mutations exhibit synergy on the Sisi phenotype in 50 mM NaI.</p

    Depletion of wildtype <i>csnk-1</i> transcripts in L4 <i>csnk-1(lf)</i> mutants.

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    (A) Positions and sequences of PCR primers for detecting all or wildtype-only csnk-1 transcripts. Partial wildtype and mutant csnk-1 sequences are aligned to show the specificity of the primers for wildtype-only transcripts. (B) Relative total csnk-1 transcript levels. (C, D) Relative wildtype-only csnk-1 transcript levels in wildtype, csnk-1(mac494lf) or csnk-1(mac495lf) animals. tba-1 was the loading control. Statistics: two-tailed unpaired Student’s t-test. *: p p p (TIFF)</p

    Representative morphologies of <i>csnk-1(lf)</i>, <i>bli-3(lf)</i> and <i>doxa-1(lf)</i> single mutants.

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    Representative morphologies of csnk-1(lf), bli-3(lf) and doxa-1(lf) single mutants.</p

    Characterization of <i>csnk-1</i>.

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    (A) csnk-1 gene structure (based on wormbase.org). The positions of mac397, mac494 and mac495 mutations are indicated. The position of the sgRNA used for CRISPR/Cas9-based mutagenesis is shown as a red bar. (B) Percentage of L1 larva that grew into adults on plates with 5 mM NaI. Results were based on two biological replicates. 100 L1 larva were analyzed in each replicate. Statistics: two-tailed unpaired Student’s t-test. *: p csnk-1p::GFP transgene in adults. (C) Fluorescent picture of a transgenic adult. The head, tail and vulva are indicated. (D-H) Higher-resolution pictures of transgenic adults. Cells with obvious GFP expression are indicated.</p

    Effects of <i>csnk-1</i> mutations or variable feeding RNAis on the Sisi phenotype.

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    Mammalian orthologs or homologs are indicated in the parentheses. (TIFF)</p

    Representative morphologies of double homozygous mutants between <i>csnk-1(lf)</i> and <i>bli-3(lf)</i> or <i>doxa-1(lf)</i>.

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    bli-3(lf) csnk-1(lf) double homozygous mutants were derived from bli-3(lf) csnk-1(lf)/hT2 heterozygous mutants. csnk-1(lf); doxa-1(lf) double homozygous mutants were derived from csnk-1(lf)/hT2; doxa-1(lf)/hT2 heterozygous mutants. Arrows point to typical blisters. All images are of the same scale. (TIFF)</p

    CSNK-1::mCherry does not colocalize with GFP and DOXA-1::GFP does not colocalize with mCherry.

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    (A, B, C) A transgenic L3 larvae co-expressing GFP and CSNK-1::mCherry in epithelial cells. (D, E, F) A transgenic 3-fold embryo co-expressing DOXA-1::GFP and mCherry in epithelial cells. For unclear reason, DOXA-1::GFP was strongly expressed in embryos but was not visible at larval stages in these transgenic lines. We therefore observed whether DOXA-1::GFP colocalizes with mCherry in embryos. (TIFF)</p

    PCR primers for generating the listed DNA fragments.

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    PCR primers for generating the listed DNA fragments.</p

    Phenotypic analyses of <i>csnk-1(lf)</i> mutants.

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    (A) Number of all eggs laid per adult. Results were based three biological replicates, with three animals per replicate. Colors represent different replicates. Statistics: two-tailed unpaired Student’s t-test. *: p csnk-1(lf) homozygous mutants. All eggs laid by a single adult on regular NGM agar plates were examined. Results were based on three biological replicates, with three animals per replicate. Statistics: two-tailed unpaired Student’s t-test. ****: p csnk-1(lf) mutants. Arrows indicate attached cuticles. (E) Quantification of young adults (24 hrs after mid-L4 larval stage) with molting defects. Results were based on three biological replicates, with 100 animals analyzed in each replicate. Statistics: two-tailed unpaired Student’s t-test. **: p p csnk-1(lf)/+ and csnk-1(lf) mutants labeled by a DPY-7::sfGFP reporter. (H) Percentage of young adults positively stained by the nuclear dye Hoechst 33258. Results were based on three biological replicates, with 59–141 animals in each replicate. Statistics: two-tailed unpaired Student’s t-test. ****: p csnk-1(lf) L4 animals stained with DCFDA. Pictures were taken with the same exposure time of 600 ms and fluorescent intensity of each animal was measured using ImageJ. (K) Quantification of DCFDA fluorescent signals of individual L4 animals. Results were based on three biological replicates, with 21–100 animals in each replicate. Statistics: two-tailed unpaired Student’s t-test. ****: p (TIFF)</p
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