Journal: Molecular Metabolism

Article Title: Long non-coding RNA LncCplx2 regulates glucose homeostasis and pancreatic β cell function

doi: 10.1016/j.molmet.2024.101878

Figure Lengend Snippet: LncCplx2 expression is regulated by BMAL1. (A) ChIP-qPCR assays demonstrating enrichment of BMAL1 at the LncCplx2 promoter region in Min6 cells. RNA Polymerase II antibody and IgG serve as the positive and negative control, respectively (n = 3/2). Two different primer pairs ( LncCplx2 -P1 and LncCplx2 -P2) targeted the promoter of LncCplx2 were used for the PCR assay. IB analysis of BMAL1, NR1D1 and E4BP4 antibody immunoprecipitation efficiency, IgG as negative control. (B) IB analysis of BMAL1 protein level in Bmal1 KO and control Min6 cells. β-Actin was used as the loading control. (C) LncCplx2 expression was analyzed by qRT-PCR in Bmal1 KO and control Min6 cells (n = 4). (D) qRT-PCR analysis of Bmal1 and LncCplx2 expression in Min6 cells expressing two shRNAs targeting Bmal1 and scrambled shRNA as control (n = 3). (E) Effects of Bmal1 overexpression on different luciferase reporter activity driven by the LncCplx2 promoter and negative control (n = 6). All the data are shown as the mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001, using two-tailed Student's t-test and one-way ANOVA.

Article Snippet: The membranes were incubated with primary antibodies against Tubulin (Proteintech, 66031-1-Ig), DDX1 (Proteintech, 11357-1-AP), SETD8 (Abcam, ab111691), HistoneH3 (Invitrogen, PA5-16183), CPLX2 (Proteintech, 18149-1-AP), BMAL1 (Invitrogen, PA1-46118), E4BP4 (Proteintech, 11773-1-AP), NR1D1 (Proteintech, 14506-1-AP), EZH2 (Invitrogen, 36-6300), and β-actin (Sigma Aldrich, A1978), followed by the appropriate HRP-conjugated secondary antibodies (Proteintech, SA00001-2/SA00001-1), and protein expression was detected with enhanced luminescence reagents (GE Healthcare, RPN2106).

Techniques: Expressing, ChIP-qPCR, Negative Control, Immunoprecipitation, Control, Quantitative RT-PCR, shRNA, Over Expression, Luciferase, Activity Assay, Two Tailed Test

Journal: Scientific Reports

Article Title: Evaluation of molecular inhibitors of loop-mediated isothermal amplification (LAMP)

doi: 10.1038/s41598-024-55241-z

Figure Lengend Snippet: Final inhibitor concentrations in LAMP reactions.

Article Snippet: The following chemicals and kits were purchased: unmodified oligonucleotides (U2Bio, Thailand), QIAamp DNA Blood Mini Kit (Qiagen, Germany), Bst 2.0 WarmStart® DNA polymerase and its buffer (New England Biolabs), EvaGreen dye (Biotium), urea (QRëC, New Zealand), humic acid (Sigma Aldrich, cat. no. H16752), tannic acid (Sigma Aldrich, cat. no. 403040), hematin porcine solution (Sigma Aldrich, cat. no. H3281), human immunoglobulin G (Sigma Aldrich, cat. no. 14506), bile salts (Sigma Aldrich, cat. no. B8756), calcium chloride (Sigma Aldrich, cat. no. 499609) and hydroxy naphthol blue (Loba Chemie Pvt Ltd, India).

Techniques: Concentration Assay

Journal: Scientific Reports

Article Title: Evaluation of molecular inhibitors of loop-mediated isothermal amplification (LAMP)

doi: 10.1038/s41598-024-55241-z

Figure Lengend Snippet: The effect of various inhibitors (bile salts, CaCl 2 , IgG and urea) on end-point hydroxy naphthol blue (HNB) LAMP reactions. Reactions were visualised after 60 min. Successful reactions give a blue colour due to a reduction in magnesium concentration at the end of the reaction, while unsuccessful reactions are purple. The concentrations of each inhibitor are: bile salts = 0.2, 0.5, 1.0, 1.5, 2.0 and 2.5 mM; CaCl 2 = 0.2, 0.4, 0.8, 1.2 and 1.5 mM; IgG = 4, 6, 8, 10, 12 and 17.4 µM; urea = 10, 50, 150, 400, 800 and 1200 mM. Pos = no inhibitor, NTC = no template control.

Article Snippet: The following chemicals and kits were purchased: unmodified oligonucleotides (U2Bio, Thailand), QIAamp DNA Blood Mini Kit (Qiagen, Germany), Bst 2.0 WarmStart® DNA polymerase and its buffer (New England Biolabs), EvaGreen dye (Biotium), urea (QRëC, New Zealand), humic acid (Sigma Aldrich, cat. no. H16752), tannic acid (Sigma Aldrich, cat. no. 403040), hematin porcine solution (Sigma Aldrich, cat. no. H3281), human immunoglobulin G (Sigma Aldrich, cat. no. 14506), bile salts (Sigma Aldrich, cat. no. B8756), calcium chloride (Sigma Aldrich, cat. no. 499609) and hydroxy naphthol blue (Loba Chemie Pvt Ltd, India).

Techniques: Concentration Assay

Journal: Scientific Reports

Article Title: Evaluation of molecular inhibitors of loop-mediated isothermal amplification (LAMP)

doi: 10.1038/s41598-024-55241-z

Figure Lengend Snippet: The effect of various inhibitors (bile salts, CaCl , IgG and urea) on real-time LAMP product detection. ( A ) shows the signal fluorescence intensity over time. Inhibitor concentrations are colour-coded per graph, where solid lines represent the mean and the shaded areas represent the 95% confidence interval for signal (N = 2/3). Arrows indicate the trend upon increasing inhibitor concentration on the time to detection of the amplicon above the background. For tabular time to detection values see Supplementary Fig. . ( B ) shows the melting profiles of the DNA amplicons at the end of the reaction. Arrows indicate the trend upon increasing inhibitor concentration on the T m (peak maxima) of the amplicons. ( C ) amplicons run on agarose gels post-stained with ViSafe Red Gel Stain. Gels are loaded with increasing inhibitor concentrations as specified in ( A ). 0 = positive control. L = 100 bp DNA base pair ladder, NTC = no template control.

Article Snippet: The following chemicals and kits were purchased: unmodified oligonucleotides (U2Bio, Thailand), QIAamp DNA Blood Mini Kit (Qiagen, Germany), Bst 2.0 WarmStart® DNA polymerase and its buffer (New England Biolabs), EvaGreen dye (Biotium), urea (QRëC, New Zealand), humic acid (Sigma Aldrich, cat. no. H16752), tannic acid (Sigma Aldrich, cat. no. 403040), hematin porcine solution (Sigma Aldrich, cat. no. H3281), human immunoglobulin G (Sigma Aldrich, cat. no. 14506), bile salts (Sigma Aldrich, cat. no. B8756), calcium chloride (Sigma Aldrich, cat. no. 499609) and hydroxy naphthol blue (Loba Chemie Pvt Ltd, India).

Techniques: Fluorescence, Concentration Assay, Amplification, Staining, Positive Control

Journal: bioRxiv

Article Title: Circadian regulation of human immunodeficiency virus type 1 replication

doi: 10.1101/2022.11.30.518024

Figure Lengend Snippet: (a) Expression of 90 HIV-1 host proteins was analysed using the CircaDatabase and 43% of genes identified as cycling in humans. (b) BMAL1 regulated genes , REV-ERB regulated genes and RORC regulated genes were compared with host factors known to alter HIV-1 replication . (c) HOMER (Hypergeometric Optimization of Motif EnRichment tool ) was used to analyse −1kb promoter regions of HIV-1 host factors and identified gene promoters encoding E-box motifs or ROR response elements (RORE). (d) Jurkat cells were treated with GSK2981278 (40 μM) or GSK805 (20 μM) for 24 h, cells were lysed, RNA was extracted, and gene expression analysed via qPCR (mean ± S.E.M., n=4, Mann-Whitney test). (e) Gene ontology (GO) cellular components analysis where each node represents an enriched GO term. Related GO terms are connected by lines, where thickness reflects the percentage of overlapping genes. See related Supplementary Figure 6.

Article Snippet: Samples were precleared with Protein A agarose beads (Millipore), immunoprecipitated with anti-REV-ERBα (NRIDI) antibody (Proteintech, 14506-I-AP), anti-BMAL1 antibody (abcam, Ab3350) or rabbit IgG (Sigma, NI01) and precipitated with Protein A agarose beads.

Techniques: Expressing, Gene Expression, MANN-WHITNEY