Total RNA for all DMS+/− samples was extracted via standard TRIzol protocol and two rounds of standard RNA ethanol precipitation. Quality of total RNA was assessed using both a ThermoFisher Scientific NanoDrop One and an Agilent Bioanalyzer 2100. All samples had a RNA Integrity Number (RIN) of 8.5–9.9, where a 10 would indicate the highest quality total RNA with no degradation and a 1 would indicate completely degraded RNA.
Nanodrop one
Manufactured by Agilent Technologies
Sourced in United States
The NanoDrop One is a compact, single-sample microvolume UV-Vis spectrophotometer designed for nucleic acid and protein quantification. It utilizes patented sample retention technology to enable measurements from 0.5 μL samples.
Automatically generated - may contain errors
Lab products found in correlation
2100 bioanalyzer, by Agilent Technologies (4 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (3 mentions)
Nextseq 500, by Illumina (3 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (3 mentions)
Total rna purification kit, by Norgen Biotek (3 mentions)
Agilent 4200 tapestation, by Agilent Technologies (3 mentions)
Agilent bioanalyzer, by Agilent Technologies (2 mentions)
Qubit 3.0 fluorometer, by Agilent Technologies (2 mentions)
Nebnext poly a mrna magnetic isolation module, by New England Biolabs (1 mentions)
Novaseq 6000 platform, by Illumina (1 mentions)
Rneasy kit, by Qiagen (1 mentions)
Mirneasy mini kit, by Qiagen (1 mentions)
Nebnext ultra 2 directional rna kit, by New England Biolabs (1 mentions)
Pt1200 e, by Thomas Scientific (1 mentions)
Mirneasy micro kit, by Qiagen (1 mentions)
On column dnase digestion, by Qiagen (1 mentions)
Hiseq 2500, by Illumina (1 mentions)
Kapa stranded mrna seq kit, by Roche (1 mentions)
Truseq stranded mrna library prep kit, by Illumina (1 mentions)
Nebnext ultra 2 directional library prep kit, by New England Biolabs (1 mentions)
13 protocols using nanodrop one
1
Total RNA Extraction and Evaluation
2
Isolation and Purification of Yeast RNA
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Total RNA from yeast cells was isolated using hot acid phenol (50 (link),51 ). RNA concentration was measured on a Nanodrop One and RNA quality was assessed by capillary electrophoresis using a PicoRNA chip on Bioanalyzer 2100 (Agilent technologies, USA). Yeast PolyA+ mRNA fraction was prepared from total RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB, US), according to manufacturer's instructions.
3
RNA-seq of venetoclax and fedratinib in RS4;11 cells
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RS4;11 cells were treated for 24 hours in triplicate (untreated, fedratinib alone, venetoclax alone, or combination). All RNA isolation and sequencing was performed by the Gene Expression Center at the University of Wisconsin-Madison. RNA was isolated with a RNeasy kit (Qiagen) prior to a quality control check using an Agilent Bioanalyzer and NanoDrop One spectrophotometer. mRNA libraries were prepared using the TruSeq stranded mRNA kit and sequenced on an Illumina NovaSeq 6000 platform to 30 million reads per sample. Sample reads were processed by the University of Wisconsin Biotechnology Center.
4
RNA Extraction from Rat Samples
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The samples of rats were randomly collected from each group in triplicate. RNA was extracted using the miRNeasy Mini Kit (Qiagen, #74106, Germany) in strict accordance with the manufacturer’s instructions. The obtained total RNA was quality-checked with an Agilent Bioanalyzer 2100 (Agilent technologies, USA) and quantified with a Qubit®3.0 Fluorometer and NanoDrop One spectrophotometer.
5
RNA-seq Protocol for Frozen Tumor Analysis
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The 27 RNA-seq datasets analyzed in this study were generated previously (5, 33 (link)). Frozen tumors underwent physical dissociation using a polytron PT1200 E homogenizer (Thomas Scientific) and total RNA was isolated using the Total RNA Purification Kit (Norgen Biotek) per manufacturer's instructions. RNA purity was quantified with the Nanodrop 2000 (Thermo Fisher Scientific) or Nanodrop One and RNA integrity was quantified with the Agilent 4200 Tapestation (Agilent Technologies). Libraries were prepared by the Cornell Transcriptional Regulation and Expression (TREx) Facility using the NEBNext Ultra II Directional RNA kit (New England Biolabs, E7760). Sequencing was performed at the Genomics Facility in the Biotechnology Research Center at Cornell University (Ithaca, NY) on the NextSeq500 (Illumina).
6
RNA-Seq Data Generation and Analysis
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Of the 27 RNA-Seq data sets analyzed in this study, 18 were generated and published previously (21 (link)), and 9 were newly generated. For the newly generated data, total RNA was isolated using the Total RNA Purification Kit (Norgen Biotek) per manufacturer’s instructions. RNA purity was quantified with the Nanodrop 2000 (Thermo Fisher Scientific) or Nanodrop One, and RNA integrity was quantified with the Agilent 4200 Tapestation (Agilent Technologies). Libraries were prepared by the Cornell Transcriptional Regulation and Expression (TREx) Facility using the NEBNext Ultra II Directional RNA kit. Sequencing was performed at the Biotechnology Research Center at Cornell University on the NextSeq500 (Illumina).
7
RNA-Seq Analysis of XRN1 Knockdown
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RNA was extracted from cell pellets, six replicates from siScrambled or siXRN1 treated cells were collected for sequencing over consecutive weeks. Total RNA was extracted using miRNEasy Micro Kit (Qiagen #217084) with on-column DNase digestion (Qiagen #79254). Total RNA concentration and quality were measured on a NanoDrop One, RNA integrity was assessed on an Agilent 2100 Bioanalyzer. RNA concentration was further assessed on a Qubit (Invitrogen #Q32852). An amount of 500 ng of total RNA was depleted for rRNA by Leeds Genomics using the Ribo-Zero Kit. Library preparation was also performed by Leeds Genomics using the Illumina TruSeq standard protocol. Subsequent libraries were run in a 75 bp single-end sequencing run on a Next Seq generating between 36 and 45 million reads per sample. Raw sequencing reads are deposited in ArrayExpress under accession number E-MTAB-10808.
8
RNA Extraction and Sequencing Protocol
9
Bacterial Nucleic Acid Quantification
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Total nucleic acid from bacterial cells, whole blood spiked with bacteria, or bacterial nucleic acids were extracted using an Allprep mini DNA/RNA kit (Qiagen, Hilden, Germany) and quantitated by NanoDrop One (Wilmington, DE) or Bioanalyzer 2100 (Agilent, Santa Clara, CA). Bacterial nucleic acid and genome equivalents were quantitated by agent-specific quantitative TaqMan real-time PCR.
10
RNA Characterization and Storage
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Isolated RNA was analyzed by NanoDrop One™, Bioanalyzer (Agilent RNA 6000 Pico Kit), and Qubit® (Qubit™ microRNA assay kit) to quantify RNA concentration and for validation of RNA quality, before stored at −80°C until further downstream analysis.
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