Nanodrop 8000

Lens ervoides elongation factor (LcEF1-α) and the A. lentis β-tubulin (TL-1) were amplified from DNA extracted from the two RILs infected with A. lentis using gene-specific primers (Supplementary Table S1). DNA was extracted with the SDS method (Goldenberger et al., 1995 (link)), quantified with a spectrophotometer (NanoDrop^TM 8000, Thermo Scientific, Waltham, United States) and adjusted to a concentration of 25 ng μl^–1. Each qPCR reaction contained 2 μl DNA template, 5 μl SYBR^® Green (Thermo Scientific), 0.2 μl of each 10 μM forward and reverse primers, and 2.6 μl distilled water. The qPCR amplifications were performed in a QuantStudio^TM 3 System (Applied Biosystems Inc., Foster City, CA, United States) using a fast-run program with default settings. The relative fungal biomass was calculated as the ratio of gDNA amplified by the TL-1 primer over that amplified by the LcEF1-α primer using the formula gDNA_F = 2^{−(CT_TL−1−CT_LCEF_1−α)}, according to the criteria used by Horevaj et al. (2011) (link).

To measure aggregation of Aβ, we modified the thioflavin T based assay described in^{69 (link)}. 500 µg Aβ42 peptide (generous gift of B. Penke, Szeged, Ungary) was dissolved and incubated for 24 h in 1 mL hexafluoro-2-propanol (HFIP) at 4 °C. Monomerized peptides were then dried via SpeedVac (Savant DNA110, Thermo Fisher Scientific, Waltham, MA, USA) at room temperature. Afterwards dried peptides were resolved in 100 µl ammonium hydroxide, sonicated for 30 min on ice, and filtered via centrifugation for 90 min at 13,900 rpm and 8 °C in Amicon Ultra-0.5 mL Filters. Final Aβ concentrations were detected via UV-Vis-spectrophotometer (NanoDropTM 8000, Thermo Fischer Scientific, Waltham, MA, USA) absorbance measurement at 280 nm. 10 µM of freshly monomerized Aβ42 was incubated with 15 µM thioflavin T on a black 96 well quartz microplate (Hellma^TM, Müllheim, Germany) in 100 µl total volume per well and in the presence of 10 µM natural compound capsaicin or ethanol as solvent control, both conditions were performed in presence of 10 µM phosphatidylcholine 16:0. Measurement was performed with an infinite M1000 Pro (Tecan, Crailsheim, Germany) at 37 °C for at least 3 days with the following wavelengths: excitation at 450 ± 5 nm and fluorescence detection at 482 ± 5 nm.

A total of 74 salivary gland tumors including 24 salivary gland adenoid cystic cancer, 28 mucoepiderrmoid cancer and 22 adenocarcinoma samples were excised from human patients under approved IRB protocols from University of Texas MD Anderson Cancer Center. MYB-NFIB fusion positive tumors were preselected by RT-PCR. Total RNAs were extracted and evaluated using NanodropTM 8000 (ThermoFisher Scientific, MA) and the Agilent 2100 Bioanalyzer with RNA 6000 Nano Labchip® (Agilent Technologies, CA).

According to the manufacturer’s instructions, RNA was isolated from cultured PHT cells at 90 h in culture using TRIzol Reagent (Invitrogen, Carlsbad, CA). RNA was resuspended in 100 μl DEPC-treated water. RNA quality was determined using an Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA), and RNA concentrations were confirmed by quantitation using a NanoDrop^TM 8000 spectrophotometer (Thermo Fisher Scientific, Wilmington, DE).

DNA was extracted from blood samples (approximately 10 drops) obtained from infants during neonatal screening using the commercial Wizard Genomic DNA purification kit (Promega, Madison, WI, USA), following the manufacturer’s procedures. DNA quality and quantity were also verified by spectrophotometry (UV–Vis) with the NanoDropTM 8000 (Thermo Fisher, Wilmington, DE, USA).

For PCR analysis, the cells were divided into NC, PC, and DLEE treatment groups. Total RNA was isolated using the Total RNA Kit according to the manufacturer’s instructions (Omega Laboratories, Inc., GA, USA). The cDNA was reverse-transcribed using a Prime Script™ cDNA Synthesis Kit (Takara, Kyoto, Japan) and quantified using a NanoDrop^TM8000 (Thermo Fisher Scientific) with Premix EX Taq™ (Takara, Kyoto, Japan). The RT-PCR process was carried out with the following conditions: denaturation for 15 s at 95 °C, annealing for 15 s at 56 °C, and extension for 30 s at 72 °C. The gene expressions were calculated using the ∆∆Ct method with glyceraldehydes-3-phosphate dehydrogenase (GAPDH) as the reference gene. Primer sequences were synthesized by Genscript as shown in Supplementary Table S1, and the fold changes were calculated with respect to the control group.