Nanodrop one uv spectrophotometer

Whole organs were homogenized in the RNA lysis buffer (Zymo Research, Irvine, CA, USA) using a shaking homogenizer with ceramic beads (MP Biomedical INC, Illkirch, France). Upon 5 min centrifugation at 10.000 g the clean supernatant was used for RNA extraction. For adipose tissue, the upper phase containing fat was discarded and not used. Upon extraction, the concentration of RNA was measured with NanoDrop One UV Spectrophotometer (ThermoFisher Scientific, Waltham, MA, USA). 2 µg of RNA was converted to cDNA using High-Capacity cDNA Reverse Transcription kit with RNase Inhibitor (Applied Biosystems, Waltham, MA, USA). The cDNA was finally diluted to a concentration of 5 ng/μL. The SsoFast EVAGreen Supermix Mastermix was used to quantify the investigated genes. For the gene expression analysis, 10 ng of cDNA was pipetted to the Mastermix. The Cycle program in the Quant 5 was 5’ @ 95 °C then 40 cycles with following conditions: 15”@95 °C and 30”@60 °C. To ensure the absence of unspecific product, a melting curve analysis between 55 °C and 95 °C was also performed at the end. Quantification of gene expression was adjusted using the Tbp mouse gene expression for the investigated adipose tissues and using Gapdh as housekeeping gene for all other tissues.

PtSMXL gene expression profiles were analyzed in roots, young leaves, mature leaves, young stems, mature stems, and dormant axillary buds of 84 K poplar. In addition, the response of PtSMXL genes to GR24 was studied in seedlings treated with six concentrations (0, 2, 5, 10, 15, or 20 μM) of GR24, and PtSMXL expression profiles during axillary bud growth were examined in axillary buds at six different stages after the begins of axillary bud growth (0, 1, 3, 5, 7, and 10 days). The more detailed description of materials is presented in the Plant materials section. Total RNA was isolated from each sample using a SteadyPure Plant RNA Extraction Kit AG21019 (Accurate Biotechnology [Hunan] Co., Ltd) and then quality-checked on a NanoDrop One UV spectrophotometer (Thermo Scientific, USA). First-strand cDNA synthesis was carried out with DNA-free RNA using Evo M-MLV Plus 1st Strand cDNA Synthesis Kit AG11615 (AG). Real-time quantitative PCR (qRT-PCR) was performed using a SYBR Green Premix Pro Taq HS qPCR Kit AG11701 (AG) on a CFX-96 real-time PCR detection system (Bio-Rad, USA). Each experiment consisted of three independent biological replicates, with three technical replicates per sample. The β-actin gene was used as an internal control. Relative expression levels of each target gene were analyzed the 2^-∆∆CT method [41 (link)]. Sequences of all primers used are listed in Table S6.

Protein elicitor samples used in the following assays were purified from the supernatant of BU412 culture through ion-exchange and size exclusion chromatography following the methods mentioned above. All protein concentrations were measured using NanoDrop One UV Spectrophotometer (Thermo Scientific).

Using an open source program, caDNAno^{53 (link)}, all structures used here were designed on the honeycomb lattice with a M13mp18 scaffold strand (7249-nt-long, GUILD, www.guildbioscience.com). Sequences of staple strands for the structures were exported from the caDNAno (Supplementary Tables 1–5) and they were synthesized from Bioneer (www.bioneer.co.kr). A folding mixture consists of 20 nM concentration of scaffold DNA, 100 nM concentration of each staple strand, 1 × TAE buffer (40 mM Tris-acetate and 1 mM EDTA, Bioneer) and 20 mM of MgCl₂ (Sigma-Aldrich, www.sigmaaldrich.com). The annealing process for self-assembly of DNA strands was performed by establishing temperature gradients from 80 to 60 °C with a rate of −0.25 °C/min and from 60 to 45 °C at a rate of −1 °C/hr in a thermocycler (T100, Bio-Rad, www.bio-rad.com). Excessive staple strands were removed through five buffer exchange procedures^{35 (link)} at 5 krcf during 8 min and concentration of structures was adjusted using the same buffer used in folding (1 × TAE and 20 mM of MgCl₂). Concentrations of folded structures were measured using a Nanodrop One UV spectrophotometer (Thermo Fisher Scientific, www.thermofisher.com). Purified structures were stored at −4 °C in a refrigerator.