guide rna lentiviral expression vector  (Addgene inc)

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: Identification of CLIP2 as a protein partner of AQP5. A , B Tandem mass spectra of CLIP2 peptide 634–645, ATLNSGPGAQQK, (+ 2 charged ion, m/z 586.4) from AQP5 IP samples in mouse parotid ( A ) and submandibular SG ( B ). Sequence specific b- and y-ions are labeled. C AQP5 immunoprecipitation (IP) performed using proteins from mouse SGs followed by WB detection using anti-CLIP2 antibodies. Input proteins from mouse submandibular glands (mSMG; lane A) and mouse parotid gland (mPG, lane B); IP performed using mSMG (lanes C, E) and mPG (lanes D, F) proteins in the presence (lanes C, D) or absence (negative controls; lanes E, F) of anti-AQP5 antibody; IP performed in the absence of any input proteins (additional negative control, lane G). D AQP5 IP performed using proteins from NS-SV-AC cells expressing AQP5 followed by WB detection using anti-CLIP2 antibodies. Input proteins from NS-SV-AC cells transfected with HA-hAQP5 (lane A) or SNAP-hAQP5 (lane B); IP performed using NS-SV-AC HA-AQP5 (lanes C, D) and NS-SV-AC SNAP-hAQP5 (lanes E, F) in the presence (lanes C, E) or in the absence (negative controls; lanes D, F) of anti-AQP5 antibody; IP performed in the absence of input proteins (additional negative control, lane G). Considering known batch-to-batch variation of commercial Sepharose-Protein A beads, whereby Protein A can shed from the beads under elution, the non-specific bands of ± 80–100 kDa ( C ) and ± 45–50 kDa ( D ) are likely non-specific bands corresponding in all likelihood to the IgG heavy chain (± 45–50 kDa) coupled or not to protein A (± 45 kDa) shed from the Sepharose beads.HA: hemagglutinin; SNAP: small protein derived from mammalian O6-alkylguanine-DNA-alkyltransferase

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: Sequencing, Labeling, Immunoprecipitation, Negative Control, Expressing, Transfection, Derivative Assay

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: Interaction between AQP5 and CLIP2 at the molecular level. A Binding curve from the MST-experiment showing the direct interaction between AQP5 and CLIP2. Data are expressed as the mean ± S.D. of bound fraction ( n = 3). The curve-line represents the curve fitting to a one-to-one binding model. B MST-data for the individual CLIP2-MTB domains (MTB1 and MTB2) showing the absence of interaction with AQP5

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: Binding Assay

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: Computer docking model of the AQP5-CLIP2 complex. A Model of the complex between the AQP5 C-terminus (orange) and the two MTB-domains of CLIP2 (light cyan and teal respectively) generated by AlphaFold Multimer. AQP5 binds primarily to MTB1 in a manner that resembles how SLAIN2 (blue) interacts with CLIP1 MTB1 (grey) and how tubulin (magenta) interacts with CLIP1 MTB2 (white). B Zoom-in on the AQP5-CLIP2 interaction site. A stretch of acidic residues (EPDED) interacts with a highly conserved basic groove on MTB1 with hydrophobic residues on the proximal (I238, Y243) and distal side (W249) interacting with hydrophobic pockets on MTB1 and MTB2 respectively. C Crystal structure of human CLIP1 MTB1 in complex with a C-terminal peptide of SLAIN2 (PDB code 3RDV) and D NMR structure of human CLIP1 MTB2 in complex with the C-terminal tail of α-tubulin (PDB code 2E4H) showing a similar mode of interaction as in the predicted AQP5-CLIP2 complex

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: Generated

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: AQP5-CLIP2 interaction and co-localization. A-B PLA showing AQP5-CLIP2 complexes in NS-SV-AC cells ( A ) expressing SNAP-AQP5 and in hMSGB from SICCA-NS and SICCA-SD patients ( B ). Arrows indicate the localization of spots. Upper-right corner inserts show representative images used for the signal quantification (scale bar:30 μm). C Quantification of AQP5-CLIP2 complexes in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the mean ± S.D. of AQP5-CLIP2 spots per cell ( n = 5). Data were analyzed using one-tailed Student’s t-test with Welch correction. D Localization of AQP5 and CLIP2 in hMSGB from SICCA-NS and SICCA-SD patients. AQP5 (AF594, red), CLIP2 (AF488, green) (scale bar: 25 μm). E Semi-quantification of AQP5 and CLIP2 localization in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the median with the interquartile range of the labelled area for each protein relative to the entire hMSGB area ( n = 3). Data were analyzed using the one-tailed Mann-Whitney U test. F Co-localization of AQP5 and CLIP2 in SICCA-NS and SICCA-SD hMSGB. Arrows indicate the AQP5-CLIP2 co-localization (yellow area). Images are shown in their original version and modified forms (used for quantification) (scale bar: 50 μm). Negative control (NEG CTRL) was conducted in the absence of primary antibodies. G Semi-quantification of AQP5 and CLIP2 co-localization in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the median with the interquartile range of the merged labelled area for both proteins relative to the entire hMSGB area ( n = 3). Data were analyzed using one-tailed Mann-Whitney U-test. One-tailed statistical tests were performed as values for SICCA-NS were not expected to be below SICCA-SD due to previously reported decreased expression of AQP5 in SICCA-SD hMSGB . C , E , G Statistical significance is indicated as *: p ≤ 0.05

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: Expressing, One-tailed Test, MANN-WHITNEY, Modification, Negative Control

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: PIP-CLIP2 interaction and co-localization. A-B PLA showing PIP-CLIP2 complexes in NS-SV-AC cells ( A ) and in hMSGB from SICCA-NS and SICCA-SD patients ( B ). Arrows indicate the localization of spots. Upper-right corner inserts show representative images used for the signal quantification (scale bar:30 μm). C Quantification of PIP-CLIP2 complexes in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the mean ± S.D. of PIP-CLIP2 spots per cell ( n = 5). Data were analyzed using one-tailed Student’s t-test with Welch correction. D Colocalization of PIP and CLIP2 in hMSGB from SICCA-NS and SICCA-SD patients. PIP (AF594, red), CLIP2 (AF488, green). Images are shown in their original version and modified forms (used for quantification) (scale bar: 50 μm). Negative control (NEG CTRL) was conducted in the absence of primary antibodies. E Semi-quantification of PIP and CLIP2 localization in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the median with the interquartile range of the labelled area for each protein relative to the entire hMSGB area ( n = 3). Data were analyzed using the one-tailed Mann-Whitney U test. C , E Statistical significance is indicated as *: p ≤ 0.05

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: One-tailed Test, Modification, Negative Control, MANN-WHITNEY

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: EZRIN-CLIP2 interaction and co-localization. A-B PLA showing Ezrin-CLIP2 complexes in NS-SV-AC cells ( A ) and in hMSGB from SICCA-NS and SICCA-SD patients ( B ). Arrows indicate the localization of spots. Upper-right corner inserts show representative images used for the signal quantification (scale bar:30 μm). C Quantification of Ezrin-CLIP2 complexes in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the mean ± S.D. of PIP-CLIP2 spots per cell ( n = 5). Data were analyzed using one-tailed Student’s t-test with Welch correction. D Colocalization of Ezrin and CLIP2 in hMSGB from SICCA-NS and SICCA-SD patients. Ezrin (AF594, red), CLIP2 (AF488, green). Images are shown in their original version and modified forms (used for quantification) (scale bar: 50 μm). Negative control (NEG CTRL) was conducted in the absence of primary antibodies. E Semi-quantification of Ezrin and CLIP2 localization in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the median with the interquartile range of the labelled area for each protein relative to the entire hMSGB area ( n = 3). Data were analyzed using the one-tailed Mann-Whitney U test. C , E Statistical significance is indicated as *: p ≤ 0.05

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: One-tailed Test, Modification, Negative Control, MANN-WHITNEY

Journal: Cell Communication and Signaling : CCS

Article Title: CLIP2: a novel functional player in AQP5 trafficking dynamics and implications for Sjögren’s disease

doi: 10.1186/s12964-025-02476-6

Figure Lengend Snippet: PIP-Ezrin interaction and co-localization. A-B PLA showing PIP-CLIP2 complexes in NS-SV-AC cells ( A ) or in hMSGB from SICCA-NS and SICCA-SD patients ( B ). Arrows indicate the localization of spots. Upper-right corner inserts show representative images used for the signal quantification (scale bar:30 μm). C Quantification of PIP-CLIP2 complexes in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the mean ± S.D. of PIP-CLIP2 spots per cell ( n = 5).Data were analyzed using one-tailed Student’s t-test with Welch correction. D Colocalization of PIP and CLIP2 in hMSGB from SICCA-NS and SICCA-SD patients. Ezrin (AF594, red), PIP (AF488, green). Images are shown in their original version and modified forms (used for quantification) (scale bar: 50 μm). Negative control (NEG CTRL) was conducted in the absence of primary antibodies. E Semi-quantification of Ezrin and CLIP2 localization in SICCA-NS and SICCA-SD hMSGB. Results are expressed as the median with the interquartile range of the labelled area for each protein relative to the entire hMSGB area ( n = 3). Data were analyzed using the one-tailed Mann-Whitney U test. C , E Statistical significance is indicated as *: p ≤ 0.05

Article Snippet: Stable transfection of NS-SV-AC cells with SNAP-hAQP5 or CRISPR/CAS9 CLIP2 dual gRNA vectors was achieved through selection with respectively 5 μg/ml of puromycin or 10 μg/ml of blasticidin (InVivo Gen, San Diego, CA, USA).

Techniques: One-tailed Test, Modification, Negative Control, MANN-WHITNEY

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A) Original and new designs of gRNA-markers for pMAGIC and nMAGIC. (B) Comparison of clone frequency in larval sensory neurons between two gRNA designs. The number represents clones between A1 and A7 segments on one side of each larva. n = larvae number: tgFE (n=10), Qtg2.1 (n=10). (C-E) Labeling of hemocytes in whole 3 rd instar larvae by pxn Gal4 >CD4-tdTom alone (C) or together with ubi-Gal80 (D) or tub-Gal80 (E). The panels on the right show enlarged views of the boxed regions. (F) Designs of Gal80 variants tested in pMAGIC gRNA-markers. (G) The brightness of epidermal clones labeled by pMAGIC gRNA-markers. n = image numbers: gRNA-40D2-uH (n = 32), gRNA-40D2-uDEH (n = 31), gRNA-42A4-uDEH (n = 52), gRNA-42A4-tDEH (n = 39), gRNA-42A4-tDES (n = 38). (H) The brightness of neuronal clones labeled by pMAGIC gRNA-markers. n = neuron numbers: gRNA-40D2-uH (n = 16), gRNA-40D2-uDEH (n = 16), gRNA-42A4-uDEH (n = 16), gRNA-42A4-tDEH (n = 15), gRNA-42A4-tDES (n = 16). (I) Portion of a larval wing disc containing nMAGIC clones visualized by nlsBFP. (J and J’) Portion of a wing disc containing nMAGIC clones labeled by cytosolic BFP (J) and HA staining (J’). (K) Epidermal clones on the larva body wall labeled by nlsBFP. (L) Epidermal clones visualized by cytosolic BFP. In all plots, black bar, mean; red bar, SD; AU, arbitrary unit. Student’s t-test in (B); one-way analysis of variance (ANOVA) and Tukey’s honest significant difference (HSD) test in (G) and (H). *p≤0.05, **p≤0.01, ***p≤0.001, ns, not significance. For (C-E), scale bar, 300 µm. For (I-M), scale bar, 100 µm.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Comparison, Clone Assay, Labeling, Staining

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A) Scheme of gRNA-marker insertion sites and target sites on Drosophila chromosomes. (B) Comparison of clone frequencies of all pMAGIC gRNA-markers in larval sensory neurons, clones are labeled using RabX4-Gal4 UAS-MApHs (for Chromosome X, II and IV) or 21-7-Gal4 UAS-MApHs (for Chromosome III). n = larvae number: X2 (n = 10), 20F2 (n = 10), 20F1(n = 10), 40D2 (n = 20), 40D4 (n = 10), 40E1 (n = 10), 41F9 (n = 20), 41F11 (n = 10), 42A4 (n = 10), 80C1 (n = 20), 80C2 (n = 14), 80F5 (n = 15), 81F (n = 10), 82A4 (n = 10), 82C3 (n = 10), 101F1a (n = 10), 101F1b (n = 10), 101F1c (n = 10). (C) Comparison of clone areas in larval wing discs labeled by nMAGIC gRNA-markers on 2R. n = wing disc number: 41F9 (n = 14), 41F11 (n = 16), 42A4 (n = 15). (D and E) Neuronal clones in the central part of the adult brain induced by pMAGIC gRNA-markers gRNA-40D2 (D) and gRNA-40E1 (E). In all plots, black bar, mean; red bar, SD. One-way ANOVA and Tukey’s HSD test. *p≤0.05, **p≤0.01, ***p≤0.001, ns, not significance. For (D) and (E), scale bar 100 µm.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Marker, Comparison, Clone Assay, Labeling

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet:

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques:

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A-F) pMAGIC clones induced in different tissues by vas-Cas9 gRNA-40D2(Gal80) and labeled by tub-Gal4 UAS-CD8-GFP (green). DAPI staining (white) shows all nuclei. (G) A pMAGIC epidermal clone on the larval body wall induced by zk-Cas9 gRNA-40D2(Gal80) and labeled by R38F11-Gal4 UAS-tdTom (green). Epidermal junctions are labeled by α-Catenin-GFP (white). (H) pMAGIC glia clones in the larval brain induced by gcm-Cas9 gRNA-40D2(Gal80) and labeled by repo-Gal4 UAS-CD8-GFP (green). Glial nuclei are labeled by Repo staining (white). (I) pMAGIC hemocyte clones induced by Act-Cas9 gRNA-40D2(Gal80) and labeled by pxn-Gal4 UAS-tdTom . For figure A, D-F, H, scale bar 100 µm. For figure B-C, G, scale bar 50 µm. For Figure I, scale bar 25 µm.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Clone Assay, Labeling, Staining

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A-A”) pMAGIC clones of VGlut 1 mutation in motor neurons at the neuromuscular junction. Clones were induced by zk-Cas9 gRNA-40D2(Gal80) and labeled by tub-Gal4 UAS-CD8-GFP ., The loss of VGlut is confirmed by VGlut staining. The mutant clones are outlined in (A”). (B-B”) A pMAGIC clone of brp d09839 mutation in a motor neuron at the neuromuscular junction. Clones were induced by zk-Cas9 gRNA-42A4(Gal80) and labeled by tub-Gal4 UAS-CD8-GFP . The loss of Brp is confirmed by Brp staining. The mutant clone is outlined in (B”). In both experiments, HRP staining shows all axons. Scale bars, 10 µm.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Clone Assay, Mutagenesis, Labeling, Staining

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A) A WT pMAGIC class IV da neuron clone exhibiting complete dendrite pruning at 16 hours APF. (B-D) pMAGIC clones of EcR M554fs mutation in da neurons imaged at 16 hours APF, exhibiting the lack of pruning (B and D) or apoptosis (C). In (A-D), the clones were induced by zk-cas9 with gRNA-41F9(Gal80) and labeled by RabX4-Gal4 UAS-MApHS . Neuronal cell bodies are indicated by arrows. MApHS contains pHluorin and tdTom , but only tdTom signals are shown. The signals in epidermal cells (A) were due to engulfment of pruned dendrites by epidermal cells . (E and F) WT (E) and Df(4)ED6380 (F) pMAGIC clones in C4da neurons induced by zk-cas9 gRNA-101Fc(Gal80) and labeled by RabX4-Gal4 UAS-MApHS . Only tdTom signals are shown. (G) Normalized dendrite length of WT clones and deficiency clones. Black bar, mean; red bar, SD. Student’s t-test. ***p≤0.001. (H) Scheme for interspecific crosses between D. melanogaster ( D.m ) and D. simulans ( D.s ). (I and J) Wing discs from male (I) and female (J) progeny carrying clones. Scale bars, 50 µm.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Clone Assay, Mutagenesis, Labeling

Journal: bioRxiv

Article Title: A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila

doi: 10.1101/2025.06.30.662354

Figure Lengend Snippet: (A-B) Representative epidermal (A) and wing disc (B) images showing uneven expression of gRNA-101F1c(BFP) inserted at attP 102D on the 4 th chromosome. Yellow arrowheads point to cells lacking BFP expression. Scale bars, 50 µm. (C) Frequency of labeled neurons by indicated gRNA(Gal80) in the absence and presence of Cas9. The zk-Cas9 dataset is the same as that for chromosome 4 in . Black bar, mean; red bar, SD. One-way ANOVA and HSD test. ***p≤0.001.

Article Snippet: The PCR product was then assembled with SapI-digested gRNA cloning vectors using NEBuilder DNA Assembly.

Techniques: Expressing, Labeling

Journal: bioRxiv

Article Title: Epitranscriptome-wide profiling identifies RNA editing events regulated by ADAR1 that are associated with DNA repair mechanisms in human TK6 cells

doi: 10.1101/2025.07.11.664482

Figure Lengend Snippet: (A) Domain structure of the two ADAR1 isoforms, p150 and p110. (B) Schematic representation of the CRISPR/Cas9-mediated disruption of ADAR1 p150 and p150/p110. The target sequence and corresponding vectors containing either a reverse-oriented puromycin- or neomycin- resistance cassette are illustrated. (C) Western blot analysis of ADAR1 isoforms. Whole-cell extracts from WT, p150 KO clones (#1, #2, #3), and p150/p110 KO clones (#1, #2, #3) were resolved by 10% SDS-PAGE. GAPDH was used as a loading control. (D) Clustering analysis of genes exhibiting A- to-I editing. Based on inosine peak scores (fold-enrichment ratio) in WT and p150 KO cells, genes were categorized as strongly p150 dependent (WT/(p150KO+1)≧4; purple), mildly p150 dependent (11, cyan). Asterisk (*) indicates the genes categorized as p150/p110 KO-specific (WT=0, p150KO=0, p150/p110KO>1).

Article Snippet: The vectors pX330-gRNA/p150 (6 μg) and the p150 target plasmid (2 μg) were transfected into TK6 cells by a NEPA21 electroporator (Nepa Gene Co. Ltd.) following the manufacturer’s instructions.

Techniques: CRISPR, Disruption, Sequencing, Western Blot, Clone Assay, SDS Page, Control

Journal: bioRxiv

Article Title: Epitranscriptome-wide profiling identifies RNA editing events regulated by ADAR1 that are associated with DNA repair mechanisms in human TK6 cells

doi: 10.1101/2025.07.11.664482

Figure Lengend Snippet: Read coverage profiles are shown for representative DNA repair-associated genes: (A) ATM (3′UTR), (B) POLH (3′UTR), (C) POLH (intron between exon4-5), (D) ATR (intron between exon1-2), (E) FANCA (intron between exon5-6), (F), FANCA (intron between exon14-15), and (G) XPA (intron between exon5-6) in wild-type (cyan), p150 KO (pink), and p150/p110 KO cells (green). The y-axis represents read counts and the x-axis indicates genomic coordinates. The scale bar corresponds to 1 kb. “Coverage data range” in the top-right corner of each panel indicates the maximum coverage value. Colored regions within coverage tracks highlight A-to-I RNA editing sites. Strand-specific colors compositions are shown: green (Adenosine) and orange (Guanosine) for forward reads; red (Thymidine) and blue (Cytidine) for reverse reads, facilitating visual estimation of nucleotide ratios at individual position.

Article Snippet: The vectors pX330-gRNA/p150 (6 μg) and the p150 target plasmid (2 μg) were transfected into TK6 cells by a NEPA21 electroporator (Nepa Gene Co. Ltd.) following the manufacturer’s instructions.

Techniques:

Journal: bioRxiv

Article Title: Epitranscriptome-wide profiling identifies RNA editing events regulated by ADAR1 that are associated with DNA repair mechanisms in human TK6 cells

doi: 10.1101/2025.07.11.664482

Figure Lengend Snippet: (A) Gene expression levels of ATM, ATR , and FANCA were quantified by RT-qPCR in WT (black), p150 KO (gray), and p150/p110 KO (white) cells. Data represent six independent experiments (n = 6). Statistical significance was determined by Student’s t-test (p < 0.05). (B) RNA-seq read coverage profiles of the XPA gene in WT#1, WT#2, p150 KO#1, p150 KO#2, p150/p110 KO#1, and p150/p110 KO#2 cells. Arrows indicate novel splicing peaks between exon 5 and 6 of the main splicing variant. (C) Detection of XPA splicing variants. PCR was performed using cDNA from WT#1, p150 KO#1, p150 KO#2, p150/p110 KO#1, and p150/p110 KO#2, with primers targeting exons 2–6 and 5–6. Amplified products were resolved on 1.0% agarose gels. For amplification between exons 2–6, expected band sizes were 946 bp for variant 1 and 1,260 bp for a potential alternative variant. For amplification between exons 5–6, expected sizes were 544 bp for variant 1 and 964 bp for alternative variants.

Article Snippet: The vectors pX330-gRNA/p150 (6 μg) and the p150 target plasmid (2 μg) were transfected into TK6 cells by a NEPA21 electroporator (Nepa Gene Co. Ltd.) following the manufacturer’s instructions.

Techniques: Gene Expression, Quantitative RT-PCR, RNA Sequencing, Variant Assay, Amplification