Journal: Communications Biology

Article Title: Loss of SIRT1 inhibits hematopoietic stem cell aging and age-dependent mixed phenotype acute leukemia

doi: 10.1038/s42003-022-03340-w

Figure Lengend Snippet: a CRISPR-mediated gene knockout of SIRT1 in human primary B/M MPAL and B-ALL cells. Indel size distribution and percentage of each indel at day 3 after electroporation were shown. There were 18% inframe indels in MPAL cells whereas inframe indels in B-ALL cells were below detection limit. b RT-qPCR analysis of SIRT1 and its target genes at day 3 after electroporation in MPAL and B-ALL cells. Wild type (WT) were mock knockout samples undergone the same procedures without sgRNAs. SIRT1 RT-qPCR primers were located at the exon 4–5 junction. c Apoptosis analysis of primary leukemia cells after SIRT1 KO at day 6. d Apoptosis analysis of two primary adult B/M MPAL samples 3 days after treatment with SIRT1 inhibitor tenovin-6 (TV-6). * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001. Error bars represented one standard deviation.

Article Snippet: The cell pellet was then resuspended in Nucleofector P3 electroporation buffer (Lonza) and then mixed with RNP solution.

Techniques: CRISPR, Gene Knockout, Electroporation, Quantitative RT-PCR, Knock-Out, Standard Deviation

Journal: Molecular Pain

Article Title: MicroRNA-9 regulates mammalian axon regeneration in peripheral nerve injury

doi: 10.1177/1744806917711612

Figure Lengend Snippet: MiR-9 overexpression inhibited axon regeneration in adult sensory neurons in vitro and in vivo. (a) qRT-PCR data indicating miR-9 levels in adult dorsal root ganglions after sciatic nerve injury. Note that miR-9 expression was significantly down-regulated from three to seven days after sciatic nerve axotomy ( n = 3 for each condition). Error bars represent SEM. ** P < 0.01. (b) and (c) Quantification of miR-9 mRNA level at three days after electroporation of miR-9 mimics in vivo and in vitro ( n = 3 for each condition). Error bars represent SEM. ** P < 0.01. (d) Representative images of EGFP-labeled regenerating axons in the whole-mount sciatic nerves. Red arrowheads mark the crush sites. Bar = 500 µm. (e) Scatter plot of average lengths of regenerating sciatic nerve axons ( n = 6 mice for each condition). Error bars represent SEM. ** P < 0.01. (f) Representative images of cultured adult mouse sensory neurons expressing EGFP (green), EGFP + miR-9 mimics (miR-9). All neurons were stained with Tuj1 (red). Scale bar = 100 µm. (g) Quantification of the average length of the longest axons (normalized to the average length of the control axons, n = 3). Error bars represent SEM. ** P < 0.01.

Article Snippet: To transfect RNA oligos into dissociated DRG neurons, the neurons were centrifuged to remove the supernatant and then resuspended in 80–100 μl of Amaxa electroporation buffer (for mouse neuron) with siRNA and miRNAs mimics (0.2 nmol per transfection) and/or the enhanced green fluorescence protein (EGFP) (10 µg per transfection) plasmid.

Techniques: Over Expression, In Vitro, In Vivo, Quantitative RT-PCR, Expressing, Electroporation, Labeling, Cell Culture, Staining

Journal: Molecular Pain

Article Title: MicroRNA-9 regulates mammalian axon regeneration in peripheral nerve injury

doi: 10.1177/1744806917711612

Figure Lengend Snippet: FoxP1 is target gene of miR-9 in adult sensory neurons during axon regeneration. (a) Representative Western blot images of FoxP1 in cultured adult dorsal root ganglion (DRG) neurons three days after transfection of miR-9 mimics. Overexpression of miR-9 leads to decreased FoxP1 protein level. (b) Quantification of FoxP1 level in vitro (normalized to actin, n = 3 for each condition). Error bars represent SEM. ** P < 0.01. (c) Representative Western blot images of FoxP1 in cultured adult DRGs in vivo three days after electroporation of miR-9 mimics. (d) Quantification of FoxP1 level in vivo (normalized to actin, n = 3 for each condition). Error bars represent SEM. ** P < 0.01.

Article Snippet: To transfect RNA oligos into dissociated DRG neurons, the neurons were centrifuged to remove the supernatant and then resuspended in 80–100 μl of Amaxa electroporation buffer (for mouse neuron) with siRNA and miRNAs mimics (0.2 nmol per transfection) and/or the enhanced green fluorescence protein (EGFP) (10 µg per transfection) plasmid.

Techniques: Western Blot, Cell Culture, Transfection, Over Expression, In Vitro, In Vivo, Electroporation

Journal:

Article Title: Experimentally Determining the IR Drop in Solution at Carbon Fiber Microelectrodes with Current Interruption and Application to Single-Cell Electroporation

doi: 10.1021/ac062045+

Figure Lengend Snippet: A) The electric field produced by the planar working electrode, on the left, is homogeneous, as shown by the evenly spaced equipotential lines. B) The electric field produced by the working microelectrode, on the right, is not homogeneous. C) The homogeneous electric field produced in bulk electroporation permits the three cells, the empty circles, to feel the same electric field. D) The inhomogeneous electric field produced by a microelectrode causes the electric field felt by the middle cell to be stronger than the other two.

Article Snippet: All chemicals were of analytical reagent grade purity and were used as received: KCl (Mallinkrodt, Paris, KT), potassium ferrocyanide, K 4 Fe(CN) 6 · 3H 2 O (Fisher Scientific, Fair Lawn, NJ), quinhydrone (Aldrich, Milwaukee, WI), methyl orange (Aldrich, Milwaukee, WI), and Iso-osmolar Electroporation Buffer (Eppendorf), which will be referred to as Iso in the rest of the paper.

Techniques: Produced, Electroporation