EMBO/sd-character-level-ner huggingface dataset:
(C) Succinate Dehydrogenase activity staining of gastrocnemius transversal sections and resulting intensity quantification (n=3-7). Data information: For all data, n represents the number of biological replicates. Statistical significance was calculated by unpaired, two-tailed Student's T-test. Data are mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 compared to control and #p < 0.05, ##p < 0.01, ###p < 0.001 compared to C26 untreated group.
Figure 8. Model of Akt-Inversin Interaction in Nephronophthisis type IIWe found that Akt interacts with and phosphorylates the ciliary proteinINVS, which is mutated in human NPHP2. Lack of phosphorylation of INVS leads to misalignment of the spindle axis and distorted luminal formation, underscoring the physiological significance of the Akt-INVS interaction in the development of abnormal cysts in NPHP2.
(C, d1) Axin2 transcripts in embryo trunk explant exposed to DMSO (C, c1, n=12/12) were reduced in Wnt-c59 exposed explants (D, d1, n=14/14). Images in (c1, d1) show TS through explants (data from 3 independent embryo explant experiments). (E, F) Slc7a5 transcripts in embryo trunk explant exposed to DMSO (E, e1, n=9/10) or Wnt-c59 (F, f1, n=15/15). Images in (e1, f1) show TS through explants (data from 3 independent embryo explant experiments). Data information: Scale bars = 100 µm
DU145 and V5-SLC1A3-DU145 cells were treated under indicated conditions with ASNase (0.2 U/ml) or TFB-TBOA (20μM) or both, and subjected to IncuCyte analysis for apoptotic cell counts.
]. (G) Quantification of quantal content. [WT n = 10 cells, hiw n = 10 cells, hiw > NADsyn-RNAi n = 10 cells. 1-way ANOVA w/Tukey's multiple comparisons, DF=29, F= 6.635, p = 0.0047. WT vs hiw p = 0.0063 (**), WT vs hiw > NADsyn-RNAi p = 0.8942 (NS)].
B. Volcano plot showing that the protein contents in sera of P15-P21 Mpdz+/+ and Mpdz-/- mice are similar to each other.
D. EM reconstruction of the Elp123 sub-complex at 27 Å resolution.E. Fitting of the Elp1 CTD into the Elp123 reconstruction.F. Fitting of the Elp2 crystal structure in the difference density generated by subtracting the partial Elp123 reconstruction from the complete Elp123 reconstruction.
(A) HEK293 cells stably expressing GFP-LC3 were transfected with control (upper) or Atg-5 siRNA (lower). After 36 h, α-synuclein fibrils were introduced into them for 4 h, followed by immunostaining with anti-phospho-α-synuclein antibody (P-αSyn). Blue, DAPI. Scale bar, 10 µm.
(d) Bif-1−/− MEFs stably transfected with a Bif-1 expression plasmid or control empty vector were cultured in EBSS for 0, 3 or 6 h and subjected to immunoblot analysis. The immunoblotting results of LC3 in c and d were quantified as the percentage of LC3-II out of total LC3 (LC3-I + LC3-II) and are represented in bar graphs.
(D) Histogram shows genome-wide distribution of log10-transformed directionality scores at bidirectional enhancers. Directionality score for enhancers is defined as the ratio of reads mapped to the plus and minus strand. Only bidirectional intergenic enhancer regions shorter than 3 kbp were considered In accordance with (Jin et al., 2017) enhancer regions were categorized as having a preferred direction (white) if the absolute value of the plus to minus strand ratio was ≥ 3 and as bidirectionally balanced (grey) if it was <3. Note that the distribution would be less balanced if unidirectionally transcribed enhancers were included.
G Touch-evoked escape responses of 48 hpf zebrafish embryos injected with ALS-linked PDI mutants (Kabashi et al, 2011; Kabashi et al, 2009). The number of touches necessary to evoke an escape response (top) and speed (bottom, in mm/s) of the escape response was determined for each condition. The total number of analyzed embryos is indicated in parenthesis. Experiments of D-F were performed in selected animals sharing normal overall embryo morphology and viability, to avoid unspecific effects of axial shortening or curvature in motoneuron morphology and/or animal behavior. Statistical analyses were performed using one-way ANOVA and Bonferroni´s post hoc tests. Mean ± SEM with only statistically significant p values are shown: *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001. Abbreviations: A (anterior), D (dorsal), P (posterior), V (ventral). Scale bars represent 500 m (C and D: a,d.g,j), 200 m (C and D: b,e,h,k), 100 m (C and D: c,f,i,l), and 25 µm (G-H).
E Correlations of somatic insertion sites of the three most represented TE families (rover, copia and I-element) with DamID tracks for adult fly intestinal stem cells (ISC).
E The immobility time in the TST of mice injected with LV- siCD39 (n = 11, 19, 15 mice. Treatment F 2, 42 = 5.807, P = 0.0059; CSDS - LV-GFP vs Ctrl - LV-GFP, P = 0.0069; CSDS - LV-siCD39 vs CSDS - LV-GFP, P = 0.0065. One-way ANOVA with Fisher's LSD test).
Venn-diagram summarizing the RNA-seq results: RNA samples were collected from V. cholerae ΔvrrA ΔmicV strains carrying pBAD-micV, pBAD-vrrA or an empty vector control. Depicted are genes displaying a fold-change of ≥3 and FDR-adjusted p-value ≤1E-8 obtained from MicV-expressing conditions (blue) or vrrA-expressing conditions (green). Genes regulated by both sRNAs (fold-change ≥3 in one condition, fold-change ≥ 2.0 in the other) are depicted in light-green.
(g) Immunoblot analysis of LAMP-2A, Itch and RCAN1 in total lysates of CD4+ T cells obtained from L2A-cKO mice (n = 3 (KO1, KO2 and KO3)) and their control littermates (n = 3) and activated for 24 h as in e,f. *, nonspecific band.
(D) HEK293T cells were transfected with SFB control, SFB- PPP2CA, SFB-PPP2CB, SFB-PPP2R1A, SFB-PPP2R1B and SFB- B56δ, and the lysates were pulled down using streptavidin beads and the interaction of endogenous PPM1G was detected by western blot using PPM1G antibody.
(D) qRT-PCR analysis of Mapk12 (p38gamma) and Mapk13 (p38delta) mRNA expression in liver extracts prepared from wild-type mice (WT) fed a diet deficient in methionine and choline (MCD) or control diet (ND) for 3 weeks; mRNA expression was normalized to the amount of Gapdh mRNA. (n=5-10)
E Immunoblots of the expression of NAD synthesis and consuming proteins Quantification values are presented as mean ± SD of four controls versus five DC samples as shown in D. Student's t test was performed on DC cells vs controls. Con: control fibroblasts.
F Population composition in male HD or patient derived untreated (n=9 HD UT, 1 Pt UT) or bulk edited T cells (Treat) from (D). For each subpopulation, the comparisons between HD Treated vs UT were performed by using LME models, with random effects defined to account for the same donor and for different number of replicates per donor within group. All p-values were adjusted with Bonferroni's correction to account for multiple testing. In all LME models, the percentages were used in square root scale to meet the assumption of normality of the residuals of the model. Mean ± SEM.
Representative western blots after subcellular protein fractionation experiment in isolated murine-derived non-tumor (NT) and GBM tumor (T) tissues.
(A) Immunofluorescent staining of C/EBP β (red) and AEP (green), Iba-1 (red) and AEP (green), cleaved Tau N368 (red) and AT8 (green), cleaved APP C856 (red) and Aβ (green), T22 (red) and AT8 (green) in cerebral cortex of brains from DSS-treated mice. Scale bar: 20 μm. White arrows indicate that C/EBP β and AEP were co-localized in mouse brain.
(E-H) Repeated pup exposure does not induce major components of social attachment behavior in Shank2-/- females: (E) pup grooming, unpaired, two-tailed Student's t-test, ***P<0.001 (F) crouching, Mann-Whitney test, ***P<0.001 and (G) nest building, Mann-Whitney test, ***P<0.001, (H) maternal interaction, unpaired, two-tailed Student's t-test, ***P<0.001 (trial 5), Shank2+/+ n = 9, Shank2-/- n = 9. Data information: All data are presented as mean ± SEM, NS: not significant.
A. Heat map of proteins identified by LC-MS/MS analysis as differentially secreted in CM of normal fibroblasts (P-NF#1 and P-NF#2), CAFs with low level of hMENA∆v6 expression (P-CAFs low#44 and #49), CAFs with high level of hMENA∆v6 expression (P-CAFs high #67 and #36). The boxed hMENA∆v6 associated signature represents proteins exclusively present in the CM derived from CAFs high. Color code is shown in the upper left corner. Co-variance = 0.2, q value = 0.1 (10% false discovery rate), adjusted P value = 0.0074542. N = 36. The values of mass spec identified for each protein were plotted in log2 scale.
A, Representative images of immunofluorescent analyses of USP33 (red) and HIF2α (green) in human primary GBMs. Frozen sections of human GBMs were immunostained with antibodies against USP33 and HIF2α, and counterstained with DAPI to show nuclei (blue). Scale bar, 40 μm. (n = 5 sections; mean ± s.e.m.).
In D, the results are expressed as a percentage of the Input for PRP8a RIP followed by RT-qPCR, and IgG RIP was used as a negative control.
E) and F) Competition assay in HeLa cells stably expressing RFP-tagged B56 inhibitor (LxxIxE) or control inhibitor (AxxAxA). YFP-B56α was transfected into and subsequently purified from these cell lines. Loss of binding of indicated proteins determined by either mass spectrometry (pink - B56 SLIM containing protein and known B56 interactor; blue - known B56 interactor, green - B56 SLIM containing protein) (E) or Western blotting (F).
(B) No correlation between the median cell size of different Ret+ subsets and eGFP intensity or IB4 binding. The graph shows the forward scatter values plotted against the normalized number of cells, displayed as the percent of Max. (C−D) Variations in endogenous eGFP intensity are clearly visible with fluorescent microscopy.
(B) Detection of WASH gene targeting. The floxed WASH gene by gene targeting was analysed by Southern blotting (left panel) and PCR (middle panel). The genotypes of the offspring were analysed by PCR (right panel).
Western blot analysis using WT and Ucp1∆L BAT lysates. The protein level of UCP1 was normalized to GAPDH (4 mo old, n = 4 per group). **p < 0.01, Student's t test
(A) Schematic of the knock-in strategy used to generate the pax2amKO2 line. Double strand break was induced between the penultimate codon and the STOP codon of pax2a gene using CRISPR/Cas9. DNA repair integrates the donor construct at the site of double strand break, resulting in a pax2a reporter line. The donor construct contains T2A-mKO2 reporter cassette flanked by left homology (LH) and right homology (RH) arms.
Expressing Tau-k18(-) or Tau-2CA abolished its anti-apoptotic function. ; N2a cells, transfected with myc-Vec, or myc-Tau40, or myc-Tau-k18(-), or myc-Tau-2CA for 48 h, were then treated with 0.5 μM STP for 4 h followed by flow cytometry using Annexin-FITC (C) Data information: Data are expressed as mean ± SEM, *, P<0.05; **, P<0.01, ***, P<0.001 vs Tau plus STP Data were analyzed by one-way ANOVA.
G) Overall Survival Prognostic value of ROCK2 expression in colon cancer patients of the GSE33582 data set (n=477 patients with survival annotation) using Kaplan-Meier survival analysis. The patient cohort was divided into tumours with or without a down-regulation of ROCK2 relatively to non-tumoural tissues (expression cut-off value corresponds to the 1st decile of ROCK2 expression in non-tumoural sample, i.e. 7.6). Log-rank test p-values for survival distribution difference are indicated at 3 and 5 years.
The rate of glucose uptake (a,e), lactate released after 4 h of incubation (b,f), glycolytic flux (c,g) and pentose-phosphate flux (d,h) were assessed in MEF and mouse cortical neurons in primary culture, as indicated, obtained from either WT or Pink1 KO (Pink1−/−) mice.
(C) The protein levels of caspase-1 p20 and IL-1β (p17) in the supernatant of cultured primary microglia were determined by western blot. (D) Quantitative analysis for Western blot analysis of caspapse-1 and IL-1β. *P < 0.05, **P<0.01 compared with the untreated group, #P < 0.05, ##P < 0.01 compared with the LPS+poly(dA:dT) group.
C) ADAR1-knockout cells (clone 1) were transfected as in (B). Protein lysates were analyzed by SDS-PAGE followed by immunoblotting using the indicated antibodies (n=4).
(I) Mobile fractions from the normalized recovery curves of NuMA-mClover at centrosomal spindle poles (control, 2-centrosomes) or at acentrosomal spindle poles (0-centrosome). Mobile fractions (roughly corresponds to normalized fluorescence intensities at the last time point were determined by single exponential fitting. Line and error bars represent the median and interquartile range (N=20 cells for each condition). The Mann-Whitney U test (two-tailed) was used to obtain a P value.
C Proportion of transcripts with extended 3´UTRs (at least 2 kb beyond the canonical polyadenylation site) among the top 100 Gadd45α hits.
C) Reads from the mRNA-seq of the SH-SY5Y neuroblastoma cells mapped to the FUS gene. The first four exons and the intervening introns of the FUS gene are depicted in purple (boxes indicate exons, thin lines indicate introns; for better visualization, intron lengths are reduced by a factor of five compared to exons). The mapped reads to this locus are shown on a log10 scale for the wild-type cells (red), FUS KO clone A4 (green) and FUS KO clone A5 (blue). In the two FUS KO clones, hardly any reads are detected downstream of the introduced ZeoR cassette.
(A) Engraftment of CD45.2 donor cells and lineage composition following transplantation of wild-type (WT) or IFNγ receptor 1 knock out (KO) HSPCs transduced with either the control Tie2.NGFR (CTRL) or Tie2.IFNγ (IFNγ) LV (mean ± SD, each dot represents an individual mouse, WT CTRL = 7 mice, WT IFNγ = 7 mice, KO CTRL = 7 mice, KO IFNγ = 7 mice).
J-L) Representative images showing that the percentage of NBs in G1-S transition and G2/M phase are both significantly increased with significantly less cells remaining in G1 phase, quantified in (L) (n= 9, 10 brain lobes). NBs in G1 phase (Mira+, GFP+) are circled by green dashed lines; NBs in G1-S transition (Mira+, GFP-RFP-) are circled by grey dashed lines; NBs in S phase (Mira+, RFP+) are circled by red dashed lines; and NBs in G2/M phase (Mira+, GFP+RFP+) are circled by yellow dashed lines.
(B) Confocal images of synaptic immunostaining in CA1 striatum lacunosum-moleculare (str. lac/mol) at P10. Arrows pointing to the enlarged synapse in the insets. (C, D) Quantification of synapse density in CA1 str. lac/mol in iCKO versus control at P10 (C), and CKO versus control at P21 (D). At P10, N(Ctrl) = 4, N(iCKO) = 3, p = 0.02. At P21, N = 8, p = 0.005.
(D-F) Immunohistological analysis of splenic sections stained with mAb against MOMA1/CD169 (blue), GFP/EYFP (green), whose expression correlates with that of IFN-β, and MCMV IE-1 (red). Scale bars, 100µm. The image shown is one representative of 15 obtained from 5 different mice. Optical magnification was 20x, and images were scanned on the microscope at zoom x2 for (E, F). White dotted lines delimit marginal zone. Yellow squares in (D) defines zoomed region in (E, F).
A Enzymatic activity of ubiquinone-cytochrome c oxidase in mitochondria isolated from cardiac tissue. The ubiquinone analogue decylubiquinone was used in this assay. Data were standardized to malate-dehydrogenase activity and presented as percent of wild-type, n= 4 per genotype.
A. In response to a 1 s light pulse, cac alleles lack on transients at day 3 and day 33.
(F) Relative expression level of PR1 in Col-0, map4k4-1, map4k3 and map4k4-1 map4k3 mutants by qRT-PCR analysis. Values are means ± SD, n = 3 (biological replicates). Data information: Different letters indicate statistical significant base on one-way ANOVA with Tukey post-test, samples sharing letters are not significantly different, p < 0.05.
The engraftment of MOLM-13 after FST knockdown was detected by flow cytometry of human CD45 and mouse CD45.1 positive cells in recipient mouse BM aspiration at week 2 post transplantation. Data information: data were presented as mean ± SEM. *P<0.05, **P<0.01 (Student's t-test).
D. Primary cilia were present in cells actively synthesizing DNA (measured by EdU incorporation). Cells were transfected with siRNA against KV10.1, and 24 hours later were incubated in the presence of EdU for 4h before fixation. KV10.1 knockdown resulted in the presence of abundant EdU-positive and simultaneously ciliated cells, suggesting a decoupling between deciliation and cell cycle progression. Scale bar: 10µm
(B) Identification of TAB2 and TAB3 fragments interacting with BCN1 in the yeast two‐hybrid system. Blue lines depict the fragments of TAB2 or TAB3 that were found to interact with BCN1 (numbers on the right refer to the amount of yeast clones identified for each fragment). The minimal domain required for the interaction is referred to as Beclin‐binding domain (BBD).
Growth kinetics of spheres formed by control and Shp2 ko cells by counting sphere cells at day 4, 7, and 12. Error bars represent SEM (n = 3).
ChIP analysis of APEX1 recruitment to the MARCKS promoter was performed except the cells expressed control shRNA (NC) or ROCKI shRNA and treated with Pam3 for 8 h. The y-axis shows the percentage of enrichment in normalized to input. An unrelated region of the ACTB2 promoter was amplified as a control.
C Schematic of the biosynthetic pathway of PS in mammalian cells. The impaired pathways in PSA‐3 and R‐41 cells are indicated.
B. Representative image of centromere 11 (red) and centromere X (green) FISH on RPE-1 CENP-AAID CENP-B KO mutant in control (top) or IAA treated condition (bottom). Scale bar represents 5 µm. Bar graphs show automatic FISH quantification of chromosome mis-segregation in the indicated cell lines with and without IAA addition for 24 h. Error bars represent the SEM of three independent experiments (n> 200 cells per experiment). Dunnett's multiple comparisons test was used to compare conditions. Grey lines separate independent experiments. Fold changes between grey and red bars are also indicated.
(F) relative proportion per experimental group of different phyla; n= 5(RDxRDT), 5(HFDxHFDT), 4(HFDxRDT); Bacteroidetes; CI 99%, Firmicutes; CI 95%, Proteobacteria; CI 95%, Actinobacteria and Spirochaetes.
B. Quantification of GIC survival using trypan blue exclusion after 24 h and 72 h of treatment with prazosin. *P=0.0286, n=4, two-sided Mann-Whitney U-test.
Herovici staining and quantification of percentage of collagen-positive (dark blue) area per field of view. N=3 tumors, n=3-4 histological sections.
Ultrastructural analysis of lungs of rhesus macaques infected with SARS-CoV-2.Lung tissue collected on 3 dpi was analyzed by transmission electron microscopy. The alveolar interstitium is expanded by edema (E), fibrin (F) and mononuclear (M) inflammatory cells (a). Normal collagen fibers (c) and multiple virions (arrowheads) line type I pneumocytes (arrows). Boxes in (a) indicate areas enlarged in (b-d). Scale bar in (a) represents 2µm, scale bars in (b-e) represent 0.2 µm.
NEDD4L was knocked out by the CRISPR Cas9 method in NHEKs. WT and NEDD4L KO NHEKs were infected with lentivirus overexpressing NEDD4L-WT or NEDD4L-C943S mutants. Cell proliferation induced by IL-6 were detected by CCK-8 Data information: Data are shown as the mean ± s.e.m., and are representative of three independent experiments Significant differences were tested using a two-tailed Mann-Whitney U test P＜0.01, ** P＜0.01, *** P＜0.001. n=4 biological replicates.
(A) Human myeloid cells and lymphoid cells were purified from PBMCs using a cell sorter. Total RNA was isolated from these primary cells and primary AECs to perform chip hybridization and microarray analysis.
(c-j) Increase in Ace2 expression in secretory cells with smoking. Mice were daily exposed to cigarette smoke or filtered air as control for two months after which cells from whole lung suspensions were analyzed by scRNA-seq (Drop-Seq). (c,d) UMAP of scRNA-seq profiles (dots) colored by experimental group (c) or by Ace2+ cells and indicated double positive cells (d). Alveolar epithelial cells (AT1 and AT2) and airway epithelial secretory and ciliated cells are marked.
D qRT-PCR analysis of Serinc3 downregulation after conditional Snai1 loss in in vitro organoid culture. VillinCreERT2 Snai1+/+ (control) and VillinCreERT2 Snai1fl/fl SI organoids were treated for 24 (P = 0.0001) and 72 h (P = 0.006) with tamoxifen. Bars represent mean ± SD. n = 3 mice per group.
Heatmaps for cell cycle, DNA replication, and p53 target genes defined by gene ontology terms in control, Shp2 ko, and Shp2‐inhibited PyMT spheres at day 7.
YTHDF1 protein expression in intestinal tissue from 4 pairs of Apc+/+ and Apcmin/+ mice. N, normal tissue. T, tumor tissue. A, tumor adjacent tissue.
(g, h) pT202/pY204 Erk1/2 immunostaining (black) in Pald1+/+ and Pald1-/- P5 pups. IsolectinB4 (white) visualizes the entire vasculature. Scale bar: 100 μm. Quantification of pT202/pY204 Erk1/2 area (h) as in the black stippled square in (g) (400 µm from the retina rim), normalized to isolectinB4 area. Mean±SEM, unpaired t-test. n=5 litters, 14 wild type and 10 knock-out retinas per genotype. Data information: *p < 0.05, **p < 0.01, ***p < 0.001.
(h) Quantification of axon number in hippocampal neurons co-transfected with GFP, Flag-TSC1 and Flag-TSC2 WT or Flag-TSC2 mutants. Quantification was performed on 150-250 neurons from each of the 3 independent experiment and the results are represented as polarity (%). All error bars represent s.e.m. *P0.05;**P0.01 compared with GFP-only transfected control. Uncropped images of western blots are shown in Supplementary Fig. S6. Source data for the statistical analysis are shown in Supplementary Table S1.
(I) HeLa cells stably expressing GFP-LC3 treated with control, PI5P4K2A, 2B, and 2C siRNA, were pretreated with Wm for 2 hr in complete medium and then shifted in HBSS media for 2 hr (in the presence of Wm) (mean ± SEM).
C. Using the indicated subfragments of INVS for IVK, we confirmed that Akt phosphorylates 675-1065 (lane 4), but not 675-822 (lane 5), or 675-746 (lane 6) INVS.
hemoglobin content (C) decrease in Foxd1Cre::Pdgfrb+/J mice continuously. Horizontal lines are means ± SD of n = 3 per group. *: p≤0.05 compared to same wt of same time point.
(A) QTL signals for ROCKI (LINC01268) gene expression in whole blood (top, black), H3K27ac promoter signal in monocytes (middle, green), and H3K4me1 signal in monocytes (bottom, orange) in a 400 kb window around the MARCKS/ROCKI locus. The signals were strongly co-localized (Pshared=.97, Pshared=.98) suggesting they represent the same signal.
C) Representative cases show the first and fourth generation xenograft H&E staining (left) and EBER-1 ISH staining (right).
B: Double-IHC of coronal brain sections (40 μm) of a littermate (LM) and a NexCre cTKO mouse (age: 5 months) stained for Ctip2 (green, expressed in layer V/VI) and Calbindin (red, expressed in layer II/III and to some extent in layer V). Note that no gross abnormalities in cortical lamination were observed. Layers of DAPI stained cell nuclei are denoted on the right (I-VI), scale bar: 100 µm.
(F) Liquid chromatography mass spectrometry analysis of the fluorogenic substrate after incubation with purified AVs. Selected ion chromatogram corresponding to different cleavage products is displayed. Numbers in graph refer to the cleavage/amino acid site from the Aβ peptide.
Principal-component analysis of differential expressed genes between 45-day-old wildtype H9 hCOs and RGCC-KO hCOs #23 based on RNA-seq. Each plot represented an individual organoid (n=3 for each groups from 1 experiment).
A. A heatmap showing the effect of single and combinatorial drug predictions by machine learning on indicated cytokine release (left), a zoomed view of top 500 drug predictions (middle), and frequency of Ponatinib occurrence in the top 500 predictions (right). A total of 92,234 single and combinatorial drug predictions were generated from 394 drugs. Predicted cytokine release is expressed as % of DMSO control.
(D qRT-PCR expression of endogenous repeats following KAP1 knockout in HeLa Results were normalized to B2M. KAP1 expression levels were verified by qRT-PCR and Western blot. Two-tailed unpaired t tests were done and p-values are: HERVK14C_1: 0.0017, HERVK14C_2: 0.0044, SVA D VNTR: 0.0243 Clones 8, 12 and 15 (from D) were selected for mRNA-sequencing.