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Nanodrop 8000 spectrophotometer

Manufactured by Agilent Technologies
Sourced in United States

The NanoDrop 8000 Spectrophotometer is a laboratory instrument designed for the quantification and analysis of nucleic acids and proteins. It utilizes a small sample volume to measure the absorbance of a sample across a wide wavelength range. The NanoDrop 8000 provides accurate and reproducible results for a variety of sample types, including DNA, RNA, and proteins.

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16 protocols using nanodrop 8000 spectrophotometer

1

Transcriptome Profiling of Malaria Mosquito

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Stage 4 larvae and 3- to 5-day-old naïve males and virgin nonblood-fed females from homokaryotypic colonies were frozen in TRIzol (Invitrogen) for subsequent total RNA extraction. Specimens were macerated using motorized pestles and the resulting total RNA samples were purified with RNeasy Mini kit (Qiagen). Concentration, quality, and integrity of the RNA samples were assessed using a Qubit 2.0 Fluorometer, a NanoDrop 8000 Spectrophotometer, and with the RNA 6000 Pico kit in a BioAnalyzer 2100 (Agilent Technologies), respectively. Unstranded (female) and stranded (male and larvae), non-polyA, ribodepleted libraries were prepared using the TruSeq Total RNA (Illumina) and the Ribo-Zero (Epicentre) kits. cDNAs were sequenced using 100-bp paired-end reads on an Illumina HiSeq 2500. Size selection was performed using 2% agarose gels on the Sage Sciences Blue Pippin, resulting in a library insert size of ∼480 nt. Three biological replicates were sequenced per biological group, that is, a particular sample type by population combination, being 27 the total number of libraries (i.e., 3 samples × 3 populations × 3 replicates). The set of nine libraries corresponding to a particular sample type was sequenced jointly in two lanes.
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2

Efficient mRNA Production for Base Editors

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Base editor mRNA was prepared by T7 run-off in vitro transcription using a custom plasmid template encoding for a T7 promoter, a minimal 5′-UTR, the base editor reading frame, 2× HBB 3′-UTR and a 110–120 bp poly(A) sequence54 (link). The plasmid template was linearized by BbsI-HF restriction digestion (NEB, R3539) and purified by phenol–chloroform extraction (Sigma-Aldrich, P2069). mRNA IVT was performed using the NEB HiScribe T7 kit (E2040S) and co-transcriptionally capped with ARCA (3′-O-Me-m7G(5′)ppp(5′)G RNA cap analogue, NEB, S1411L) or CleanCap AG (Trilink, N-7113). Partial (75–85%) or total UTP substitution with N1-methyl-pseudo-UTP (Trilink, N-1103) or 5-methoxy-UTP (Trilink, N-1093) was performed as indicated. Dephosphorylation with QuickCIP (NEB, M0525L) or DNase treatment (NEB, M0303L) was added after IVT reaction (30 min at 37 °C). IVT mRNA was purified using the NEB Monarch RNA Clean up kit (T2050L) or sparQ PureMag magnetic beads (MagBio, 95196) and resuspended in RNase-free water. mRNA was quantified using the Nanodrop-8000 spectrophotometer and quality control was performed using the Agilent Fragment Analyzer with RNA Kit-15NT (Agilent, DNF-471).
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3

RNA-seq Analysis of Prostate Cancer Cell Lines

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GE of all prostate cancer and normal prostate cell lines at baseline (no treatment) was assessed by RNA-seq. Furthermore, the effects of METRO-TOPO and CONV-TOPO exposure for 6 weeks on ARLow mCRPC/NEPC PC-3 tumor model (3D spheroid) were assessed using RNA-seq. Pre- and post-drug exposure, as described above, tumor cells were harvested, and high-quality RNA was extracted using QIAshredder and RNeasy kit (Qiagen). RNA concentration and integrity were assessed using a Nanodrop-8000 spectrophotometer and Agilent 2100 Bioanalyzer. An RNA integrity number threshold >8 was applied, and RNA-seq libraries were constructed using Illumina TruSeq RNA Sample Preparation kit v2. Libraries were then size-selected to generate inserts of approximately 200 bp. RNA-seq was performed on Illumina's NovaSeq platform using a 150 bp paired-end protocol with a depth of >20 million reads per sample. Average quality scores were above Q30 for all libraries in both R1 and R2 (29 (link)). AA cell line RNA was isolated from cultured cells using TRIzol Reagent (Sigma Life Sciences) following the manufacturer's protocol (27 (link)). Library preparation, quality control, and sequencing of extracted RNA were performed by the Center for Pharmacogenomics and Single-Cell Omics (AUPharmGX).
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4

Illumina HumanHT-12 v4 Expression Analysis

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Total RNA samples were assessed for concentration using Thermo Scientific's NanoDrop 8000 Spectrophotometer (Carlsbad, CA) and degradation using the Agilent RNA 6000 Nano Kit with the Agilent 2100 Bioanalyzer (Santa Clara, CA). 250 ng of total RNA was amplified and biotinylated with Ambion's Illumina® TotalPrep™ RNA Amplification Kit (Austin, TX) for hybridization to Illumina HumanHT-12 v4 Expression BeadChip (La Jolla, CA) per manufacturer's instructions. Expression profiling was performed on two individual cell clones per group and profiles were deposited in the GEO repository (accession number GSE63529).
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5

RNA Isolation, RT-qPCR Quantification

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RNA was isolated with the RNeasy mini kit (Qiagen) or by the High Pure RNA isolation kit (Roche) and subsequently quantified using the NanoDrop 8000 spectrophotometer (Agilent). RNA was reversed transcribed using the Transcriptor First Strand Kit (Roche). qPCR was carried out using SYBR Green on a LightCycler 480 (Roche) with absolute quantification by generating standard curves for each primer pair. Gene expression was displayed relative to HPRT.
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6

Transcriptome Analysis of Stachys tora

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Specimens of S. tora (cv. Myeongyun) were grown in an experimental plot of National Institute of Horticultural and Herbal Science (Eumseong) field. The distance between two adjacent plants was maintained at 50×40 cm and fertilizer (N-P20-K20) was applied at 8–10 Kg per 10a. Leaf, root, and early- and late-stage seed tissues were harvested from healthy plants, and stored at -80°C until used for RNA extraction. Total RNA was extracted from leafs, roots, and two stages of seeds of S. tora using the RNeasy Plant Mini kit (Qiagen, InS., Valencia, CA, USA). RNA purity was determined using NanoDrop8000 Spectrophotometer and Agilent Technologies 2100 Bioanalyzer, and total RNA integrity was identified as having a minimum integrity value of 7.
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7

Total RNA Sequencing by Illumina

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Total-Stranded RNAseq sequencing was performed by the Centre National de Recherche en Génomique Humaine (CNRGH), Institut de Biologie François Jacob. After complete RNA quality control on each sample (quantification in duplicate on a NanoDrop 8000 spectrophotometer and RNA6000 Nano LabChip analysis on Bioanalyzer from Agilent), libraries have been prepared using the “TruSeq Stranded Total RNA” Kit from Illumina. An input of 1 μg total RNA was used for all samples, and libraries were prepared according to manufacturer’s instructions. After library quality control and quantification, sample libraries have been pooled before sequencing to reach the expected sequencing depth. Sequencing has been performed on an Illumina HiSeq200 as paired-end 100 bp reads, using Illumina sequencing reagents. Fastq files produced after RNA-seq have been be processed by in-house CNRGH tools to assess quality of raw and genomic-aligned nucleotides.
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8

Extraction and Quality Assessment of Crab DNA and RNA

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Three live adult red king crab individuals were kindly provided by Norway King Crab Ltd (https://nkc.no). They were stored in a freezer at − 80 °C before DNA and RNA isolation.
Total DNA was extracted from each sample with the DNeasy Blood & Tissue Kit (Qiagen, Cat No: 69504) according to the manufacturer’s instructions. The concentration and integrity of the RNA were assessed with a Thermo Scientific NanoDrop 8,000 Spectrophotometer and Agilent 2,100 Bioanalyzer, respectively (Agilent Technologies, USA).
RNA was extracted from four tissues from three adult individuals; see Table 2 for sample description. Total RNA was extracted from each sample with the Direct-zol RNA Miniprep Plus kit (Zymo, R2071) according to the manufacturer’s instructions after treatment with RNase-free DNase I (Qiagen) to eliminate genomic DNA. The concentration and integrity of the RNA were assessed with a Thermo Scientific NanoDrop 8,000 Spectrophotometer and Agilent 2,100 Bioanalyzer, respectively (Agilent Technologies, USA).
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9

Globin Depletion for RNA-Seq

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In order to assess the technology platform dynamic range and lower limit of detection, External RNA Controls Consortium (ERCC) RNA spike-in control mix (cat#4456740, Invitrogen, San Diego, CA, USA) was added to each RNA sample before further processing. The 12 total RNA aliquots (6 biological replicates, 6 technical replicates) destined for globin transcript depletion were further processed as follows. RNA was concentrated using RNeasy MinElute Cleanup Kit (cat#74204, Qiagen) to reduce sample volume before globin transcript depletion. Human alpha and beta globin mRNA was then depleted from total RNA using the GLOBINclear kit (cat#AM1980, Ambion- Applied Biosystems, Mississauga, ON, Canada) and following the manufacturer's protocols. Prior to sequencing, the amount and quality of all 24 RNA samples were assessed using a NanoDrop8000 Spectrophotometer and Agilent 2100 Bioanalyzer.
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10

RNA Isolation from Frozen Kidney Tissue

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The kidney tissues snap-frozen and stored at -80°C were used for mRNA microarray analyses as previously described [21 (link)]. Total RNA was isolated using the miRNeasy 96 kit (Qiagen, Cat No 217061, Valencia, CA), according to the manufacturer’s instructions. Quality and quantity of RNA were evaluated with a 2100 Bioanalyzer (Cat. No. 62947, Agilent Technologies, Santa Clara, CA), using the Agilent RNA 6000 Nano Reagents (Cat. No. 5067–1511, Agilent Technologies,), and a multi-well Nanodrop 8000 spectrophotometer (Cat. No. 1464, Scientific, Waltham, MA).
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