Journal: PLOS ONE

Article Title: Evaluating possible maternal effect lethality and genetic background effects in Naa10 knockout mice

doi: 10.1371/journal.pone.0301328

Figure Lengend Snippet: Genotypes of E9.5-E13.5 embryos from different mating series.

Article Snippet: Chi Squared Analysis for E9.5-E13.5 embryos only (Analyzing only Naa10 -/Y vs Naa10 +/- and Naa10 -/+ ) , , , , , P>0.05.

Techniques:

Journal: PLOS ONE

Article Title: Evaluating possible maternal effect lethality and genetic background effects in Naa10 knockout mice

doi: 10.1371/journal.pone.0301328

Figure Lengend Snippet: Chi squared analyses.

Article Snippet: Chi Squared Analysis for E9.5-E13.5 embryos only (Analyzing only Naa10 -/Y vs Naa10 +/- and Naa10 -/+ ) , , , , , P>0.05.

Techniques:

Journal: Cancer Science

Article Title: NeuroD1 promotes neuroblastoma cell growth by inducing the expression of ALK

doi: 10.1111/cas.12628

Figure Lengend Snippet: Relationship of NeuroD1 and ALK in neurospheres. (a) NeuroD1 in neurosphere cells cultured from the hippocampus region of wild-type E13.5 embryos was knocked down by two independent shRNA. 4 days later, the phase contrast pictures were taken. Scale bar, 200 μm. (b) The sphere numbers in (a) were counted. (c) The mRNA levels of NeuroD1 were examined by real-time PCR. (d) The mRNA levels of ALK were examined by real-time PCR. * P < 0.05. ** P < 0.01.

Article Snippet: Embryonic neural stem cells were prepared from an E13.5 ICR mouse (Japan SLC, Hamamatsu, Japan).

Techniques: Cell Culture, shRNA, Real-time Polymerase Chain Reaction

Journal: Nature neuroscience

Article Title: Balanced gene regulation by an embryonic brain non-coding RNA is critical for GABA circuitry in adult hippocampus

doi: 10.1038/nn.2371

Figure Lengend Snippet: a. Schematic of targeting a triple polyadenylation transcription stop site into Evf exon1, truncating Evf2 transcripts from 3.7 kb to 101 bp (Evf2TS), but not Evf1, which is transcribed starting from exon3. Truncated Evf2TS transcript (101 bp) completely lacks the ultraconserved ei region. Only the distance from TS insertion to the Dlx5 (~10.2 kb) and Evf1 (~10.4 kb) transcription start sites are shown to scale. b. ES cells used for making Evf2TS mice contain correctly targeted transcription stop into Evf exon1, as verified by Southern analysis. c–h. RNA in situ hybridization of E13.5 coronal sections of wildtype (wt) and Evf2 TS/TS mutant telencephalon, probed with anti-sense Evf2 (c, d), anti-sense Evf1 (e, f), or anti-sense Dlx5 (g, h). i. Quantitative real-time RT-PCR analysis of E13.5 MGE from wt and Evf2 TS/TS mutants. Error bars: S.E.M. Statistical analysis: Mann Whitney U-test, p<.05: Evf2=0.0055, Dlx5=0.044, Dlx6=0.0055; n= 4 for wt, n=6 for Evf2 TS/TS . j. Quantitative real-time RT-PCR analysis of E12.5 Evf2 TS/TS mutant MGE electroporated with: control (pcDNA, 2μg), 1 μg pcDNA-Evf2 (+1 μg pcDNA), and 2μg pcDNA-Evf2. Error bars: S.E.M. Statistical analysis, ANOVA Dunnett’s two-sided test, p<.05 values: Dlx5, (0μg,1μg Evf2=0.05, 0μg,2μg Evf2=0.001), ANOVA Tukey test, p<.05 values, Dlx6 (0μg,2μg Evf2=0.041, 1μg Evf2, 2μg Evf2= .033); n=4. * p < 0.05; t -test, Mann Whitney, or ANOVA

Article Snippet: Total RNA from pairs of E13.5 MGE tissue was isolated and treated with RNASE-free DNASE I (NEB), and reverse-transcribed using random hexamers (NEB) and MMoLV reverse-transcriptase (Invitrogen).

Techniques: RNA In Situ Hybridization, Mutagenesis, Quantitative RT-PCR, MANN-WHITNEY

Journal: Nature neuroscience

Article Title: Balanced gene regulation by an embryonic brain non-coding RNA is critical for GABA circuitry in adult hippocampus

doi: 10.1038/nn.2371

Figure Lengend Snippet: a. Schematic showing the region in mouse chromosome 6qA1 surrounding Dlx5/6 and Evf1/2. Quantitative chromatin immunoprecipitation of E13.5 MGE from wt (black bars) and Evf2 TS/TS mutants (gray bars) using primers 1–6 across the Dlx 5/6 region, and antibodies to: b. anti-pan DLX, p <.05 values: 3=0.025, 5=0.025, c. anti-MECP2, p<.05 values: 2=0.025, 3=0.025, 5=0.025, d. anti-HDAC1, p <.05 values: 5=0.025, 6=0.025. qChIP-PCR was performed on optimized primer sets using previously defined primer sets (1=15, 2=24, 4=27 ) and newly defined sets for ei (3- red), eii (5-red) and external primer (6). Primer 2 (green) was identified as a MECP2/HDAC binding site for transcriptional repression in adult cortex . Error bars: S.E.M. Statistical analysis: Mann Whitney U-test. * p < 0.05 Mann Whitney U-test. The schematic in ( a ) is reversed from that shown for Evf2 transcription-stop insertion in order to align with previously published primers .

Article Snippet: Total RNA from pairs of E13.5 MGE tissue was isolated and treated with RNASE-free DNASE I (NEB), and reverse-transcribed using random hexamers (NEB) and MMoLV reverse-transcriptase (Invitrogen).

Techniques: Chromatin Immunoprecipitation, Binding Assay, MANN-WHITNEY

Journal: Nature neuroscience

Article Title: Balanced gene regulation by an embryonic brain non-coding RNA is critical for GABA circuitry in adult hippocampus

doi: 10.1038/nn.2371

Figure Lengend Snippet: a. Western Blot analysis of DLX2 protein after transfection of a construct containing Dlx2 cDNA and/or Evf2 cDNA. Transfection was performed into C17 mouse neural stem cells in the presence or absence of Evf2 RNA, showing equal levels of DLX2 protein. That Evf2 RNA is stable after transfection was verified by RT-PCR (not shown). b. Western analysis of Evf2 TS/TS and wt E13.5 ganglionic eminence extracts probed with anti-DLX2 specific antibody (gift of David Eisentstat) shows that loss of Evf2 does not affect DLX2 protein levels. c–h. Pan-anti-DLX antibody (green) stains nuclei counterstained with DAPI (blue) in both wt and Evf2 TS/TS E13.5 MGE. MGE, medial ganglionic eminence; VZ, ventricular zone; SVZ, subventricular zone. c, f: scale bar= 200μm; d, g: scale bar=20μm; e, h: scale bar= 10μm. i–j. Immunohistochemistry using anti-MECP2 antibodies (green) and DAPI (blue nuclei) shows similar nuclear localization in both wt and Evf2 TS/TS MGE cells, scale bar=20μm.

Article Snippet: Total RNA from pairs of E13.5 MGE tissue was isolated and treated with RNASE-free DNASE I (NEB), and reverse-transcribed using random hexamers (NEB) and MMoLV reverse-transcriptase (Invitrogen).

Techniques: Western Blot, Transfection, Construct, Reverse Transcription Polymerase Chain Reaction, Immunohistochemistry

Journal: Nature neuroscience

Article Title: Balanced gene regulation by an embryonic brain non-coding RNA is critical for GABA circuitry in adult hippocampus

doi: 10.1038/nn.2371

Figure Lengend Snippet: a. Quantitative real-time RT-PCR analysis of GAD67 in E13.5 MGE from wt and Evf2 TS/TS mutants. Error bars: S.E.M. Statistical analysis: Student’s t -test, p=0.031; n= 3 for wt, n=5 for Evf2 TS/TS . b. Quantitative real-time RT-PCR analysis of GAD67 in E12.5 Evf2 TS/TS mutant MGE electroporated with: control (pcDNA, 2μg), and 2μg pcDNA-Evf2. Error bars: S.E.M. Statistical analysis: Student’s t -test, p=0.0375; n=4. c. Quantitative real-time RT-PCR analysis of GAD67 in P60 hippocampus from wt and Evf2 TS/TS mutants. Error bars: S.E.M. Statistical analysis: Student’s t -test, p=0.031; n=2 for wt, n=2 for Evf2 TS/TS . d. Quantification of the number of GAD67-expressing GABAergic interneurons in 8-month old dentate gyrus (DG), and hippocampal CA3 and CA1 regions. Error bars: S.E.M. Statistical analysis: Student’s t -test, p>0.05 for DG, CA3, CA1; n=3 for wt, n=3 for Evf2 TS/TS . * p < 0.05; t -test

Article Snippet: Total RNA from pairs of E13.5 MGE tissue was isolated and treated with RNASE-free DNASE I (NEB), and reverse-transcribed using random hexamers (NEB) and MMoLV reverse-transcriptase (Invitrogen).

Techniques: Quantitative RT-PCR, Mutagenesis, Expressing