Journal: Insects

Article Title: Evidence of the Involvement of a Plus-C Odorant-Binding Protein HparOBP14 in Host Plant Selection and Oviposition of the Scarab Beetle Holotrichia parallela

doi: 10.3390/insects12050430

Figure Lengend Snippet: Binding of different compounds to recombinant OBP14 of H. parallela .

Article Snippet: The His-tag of the purified recombinant protein was removed by using recombinant enterokinase (Shanghai Korain Biotech Co Ltd., Shanghai, China), then a second purification was performed with the Ni affinity chromatography.

Techniques: Binding Assay, Recombinant

Journal: The Journal of Biological Chemistry

Article Title: An N-terminal di-proline motif is essential for fatty acid–dependent degradation of Δ9-desaturase in Drosophila

doi: 10.1074/jbc.M117.801936

Figure Lengend Snippet: Role of the di-proline motif in degradation of Δ9-desaturase. N-terminal amino acid sequences of Δ9-desaturases were compared (D. melanogaster DESAT1 (NP_652731), Mus musculus SCD1 (NP_033153), M. musculus SCD3 (NP_077770), Homo sapiens SCD1 (NP_005054), Danio rerio stearoyl-CoA desaturase (AAO25582), and S. cerevisiae Ole1 (CAA96757)) (A). Schematic illustration of DESAT1, mouse SCD1, and constructed mutants is shown (B). S2 cells expressing mouse SCD1-FLAGC, chimera-FLAGC, chimera (AA)-FLAGC, and SCD1 (di-Pro)-FLAGC were treated with C18:1 (100 μm) for 6 h (C) or DESAT1 inhibitor 37c (1 μm) for 16 h (D), and the amounts of endogenous DESAT1, FLAG-tagged exogenous Δ9-desaturases, and α-tubulin protein were detected with anti-DESAT1 antibody, anti-FLAG antibody, and anti-α-tubulin antibody, respectively. Band intensities were determined by ImageJ software, and levels of SCD1 proteins are shown relative to the amount of SCD1 protein in vehicle-treated cells (C and D). Mean ± S.D. (n = 3). *, p < 0.05; ***, p < 0.001; n.s., not significant.

Article Snippet: Anti-α-tubulin antibody (PM054), anti-FLAG antibody (M2), and anti-ubiquitin antibody (SPC-119) were purchased from MBL, Sigma, and StressMarq Biosciences, respectively.

Techniques: Construct, Expressing, Software

Journal: The Journal of Biological Chemistry

Article Title: An N-terminal di-proline motif is essential for fatty acid–dependent degradation of Δ9-desaturase in Drosophila

doi: 10.1074/jbc.M117.801936

Figure Lengend Snippet: Identification of protease involved in the degradation of DESAT1. S2 cells were treated with calpeptin (50 μm), MG132 (50 μg/ml), or chloroquine (50 μm) for 6 h (A). S2 cells were treated with dsRNA against GFP, CalpA, CalpB, CalpC, or sol for 72 h and then exposed to C18:1 (100 μm) for 6 h (B). S2 cells expressing DESAT1-FLAGC and DESAT1 (P2A/P3A)-FLAGC were treated with calpeptin (50 μm) for 6 h (C). The amounts of endogenous DESAT1, FLAG-tagged exogenously expressed DESAT1, and α-tubulin protein were detected with anti-DESAT1 antibody, anti-FLAG antibody, and anti-α-tubulin antibody, respectively (A–C). Band intensities were determined by ImageJ software, and levels of DESAT1 proteins are shown relative to the amount of DESAT1 protein in vehicle-treated cells (A–C). Mean ± S.D. (n = 3). **, p < 0.01; ***, p < 0.001; n.s., not significant.

Article Snippet: Anti-α-tubulin antibody (PM054), anti-FLAG antibody (M2), and anti-ubiquitin antibody (SPC-119) were purchased from MBL, Sigma, and StressMarq Biosciences, respectively.

Techniques: Expressing, Software

Journal: Cell Reports Methods

Article Title: Derivation of nociceptive sensory neurons from hiPSCs with early patterning and temporally controlled NEUROG2 overexpression

doi: 10.1016/j.crmeth.2022.100341

Figure Lengend Snippet: Comparison of SN differentiation protocols with combined small-molecule patterning and overexpression of NGN2 (A) Gene expression profiles of small-molecule-only differentiations (standard) and hybrid differentiations (hybrid) were compared using qRT-PCR. In all panels, data show means ± SD normalized to housekeeping genes. Values for hybrid cultures were normalized to day 11 of standard culture values. ∗adjusted p < 0.05, ∗∗adjusted p < 0.01 as determined by Sidak’s multiple comparisons tests after two-way ANOVA. (B) Representative ICC images highlighting the expression of BRN3A and ISL1 in cultures of all lines and conditions, performed on day 35. Scale bars show 100 μm. (C) Quantification of immunohistochemistry images. Data shows means ± SD. Dissociated neurons were stained with fluorophore-conjugated antibodies for TRKA, TRKB, and TRKC. (D) Representative histograms of cytometry data for the G3_H condition, showing counts of cell fluorescence for each of the antibodies, normalized to the mode of each distribution. (E) Cytometry data were quantified to provide an estimate of the number of single cells expressing each of the TRK proteins. Data shows means ± SD. ∗adjusted p < 0.05, ∗∗adjusted p < 0.01 as determined by Sidak’s multiple comparisons tests after two-way ANOVA. Data in (A), (C), and (E) comprise results from at least 3 differentiations for each condition.

Article Snippet: The following primary antibodies were used: Sox10 (#89356, Cell Signalling) at 1:1000 dilution; NEUROG1 (#mab3524, R&D) at 1:500 dilution, BRN3A. (mab1585, Milipore) at 1:200, ISL1 (AB178400, Abcam) at 1:100 dilution, Tuj1 (MAB1637, Millipore) at 1:500 dilution, PRPH (AB4666, Abcam) at 1:100.

Techniques: Over Expression, Expressing, Quantitative RT-PCR, Immunohistochemistry, Staining, Cytometry, Fluorescence

Journal: Cell Reports Methods

Article Title: Derivation of nociceptive sensory neurons from hiPSCs with early patterning and temporally controlled NEUROG2 overexpression

doi: 10.1016/j.crmeth.2022.100341

Figure Lengend Snippet:

Article Snippet: The following primary antibodies were used: Sox10 (#89356, Cell Signalling) at 1:1000 dilution; NEUROG1 (#mab3524, R&D) at 1:500 dilution, BRN3A. (mab1585, Milipore) at 1:200, ISL1 (AB178400, Abcam) at 1:100 dilution, Tuj1 (MAB1637, Millipore) at 1:500 dilution, PRPH (AB4666, Abcam) at 1:100.

Techniques: Recombinant, Knock-Out, RNA Extraction, RNA Sequencing Assay, Software