The quality of RNA was determined by using an RNA 6000 Nano LabChip Kit and Agilent Bioanalyser 2100 (Agilent, Palo Alto, CA, USA). Preparation of cRNA was performed according to the protocol provided by Affymetrix (Santa Clara, CA). The same amounts of total RNA from two animals were pooled and further purified by using an RNeasy Mini Kit (Qiagen Inc. Total RNA (3 μg) derived from each pool (n = 4) was converted to double-stranded cDNA using a SuperScript System (Invitrogen, Carlsbad, CA) and an oligo(dT)24 primer containing a T7 RNA polymerase promoter site. Biotin-labelled cRNA was synthesised from cDNA using a labelling Kit and purified by using a GeneChip Cleanup Sample Module (Qiagen Inc., Valencia, CA, USA). The yield of the in vitro transcription reaction was determined by the product absorbance at 260 nm, as measured by NanoDrop ND-1000 (NanoDrop Technologies, Inc., Montchanin, DE), size of cRNA probes was evaluated by using RNA 6000 Nano LabChip Kit (Agilent, Palo Alto, CA, USA). Fragmented cRNA was used for hybridization to GeneChip® Rat Gene 2.0 ST arrays (Affymetrix). The arrays were washed and stained with streptavidin-phycoerythrin (Merck, Darmstadt, Germany) in Fluidics Station 400 (Affymetrix) according to the standard protocol of the manufacturer. The arrays were scanned by using a GeneChip Scanner 3000 (Affymetrix).
Rna 6000 nano labchip kit
Manufactured by Agilent Technologies
Sourced in United States, Germany, Ireland, Spain, France, Canada, China
The RNA 6000 Nano LabChip Kit is a laboratory instrument designed for the analysis and quantification of RNA samples. It utilizes microfluidic technology to assess the integrity and concentration of RNA in a small sample volume.
Automatically generated - may contain errors
Lab products found in correlation
2100 bioanalyzer, by Agilent Technologies (259 mentions)
Trizol reagent, by Thermo Fisher Scientific (210 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (158 mentions)
Mrna seq sample preparation kit, by Illumina (61 mentions)
Poly t oligo attached magnetic beads, by Thermo Fisher Scientific (53 mentions)
Rneasy mini kit, by Qiagen (48 mentions)
Nanodrop nd 1000 spectrophotometer, by Thermo Fisher Scientific (41 mentions)
Hiseq 4000, by Illumina (35 mentions)
Hiseq 2500, by Illumina (31 mentions)
Nanodrop 1000 spectrophotometer, by Thermo Fisher Scientific (31 mentions)
Trizol, by Thermo Fisher Scientific (29 mentions)
Agilent 2100 bioanalyser, by Agilent Technologies (22 mentions)
Truseq small rna sample prep kit, by Illumina (20 mentions)
Novaseq 6000, by Illumina (18 mentions)
Novaseq 6000 platform, by Illumina (18 mentions)
Hiseq 4000 platform, by Illumina (14 mentions)
Epicentre ribo zero gold kit, by Illumina (13 mentions)
Nanodrop nd 1000, by Thermo Fisher Scientific (12 mentions)
Agilent bioanalyzer 2100 system, by Agilent Technologies (12 mentions)
Rneasy plus mini kit, by Qiagen (11 mentions)
530 protocols using rna 6000 nano labchip kit
1
Agilent Bioanalyzer RNA Profiling
2
Transcriptome Analysis of Rat Gene Expression
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The quality of RNA was determined by using RNA 6000 Nano LabChip Kit and Agilent Bioanalyzer 2100 (Agilent, Palo Alto, CA, USA). Preparation of cRNA was performed according to the protocol provided by Affymetrix (Santa Clara, CA). The total RNA from individual animal was further purified by using RNeasy Mini Kit (Qiagen Inc., Valencia, CA, USA). Total RNA (3 μg) was converted to double-stranded cDNA using SuperScript System (Invitrogen, Carlsbad, CA) and an oligo (dT24) primer containing a T7 RNA polymerase promoter site. Biotin-labeled cRNA was synthesized from cDNA using a labeling kit and purified by using a GeneChip Cleanup Sample Module (Qiagen Inc., Valencia, CA, USA). The yield of the in vitro transcription reaction was determined by product absorbance at 260 nm measured by NanoDrop ND-1000 (NanoDrop Technologies, Inc., Montchanin, DE), and a size of cRNA probes was evaluated by using RNA 6000 Nano LabChip Kit (Agilent, Palo Alto, CA, USA). Fragmented cRNA was used for hybridization to GeneChip® Rat Gene 2.0 ST arrays (Affymetrix). Arrays were washed and stained with streptavidin-phycoerythrin (Merck, Darmstadt, Germany) in Fluidic Station 400 (Affymetrix), according to the standard protocol of the manufacturer. The arrays were scanned by using GeneChip Scanner 3000 (Affymetrix).
3
Profiling miRNA Expression in H. pylori Infection
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HET‐1A and OE33 cells were co‐cultured with or without H. pylori 26695 for 12 hours. The total RNA of the cells was isolated using TRIzol reagent (Invitrogen, Carlsbad, CA, USA), and the miRNA fraction was further purified using a mirVana miRNA isolation kit (Ambion, Austen, TX, USA). For microarray analysis, the quality of RNA and the RNA integrity number were determined by using RNA 6000 Nano LabChip Kits (Agilent Technologies, Santa Clara, CA, USA). An Agilent Human miRNA microarray chip version, containing 4947 probes corresponding to 1887 unique human miRNAs and 140 human viral miRNAs cataloged in the Sanger database version 18.0 (Agilent Technologies), was used according to the manufacturer's recommendations. The difference in miRNA expression between the cells infected with or without H. pylori was considered significant if the change of expression was more than 1.5‐fold and the t‐test P‐values were <.05. Target gene prediction of the screened miRNA was carried out using the online miRNA target predicting software TargetScan (www.targetscan.org ), miRBase (www.mirbase.org ), and miRanda (www.microrna.org ). Only genes predicted by at least two software programs were considered as potential target genes of the miRNA.
4
Transcriptomic Analysis of RNA Samples
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Total RNAs were extracted using the GenEluteTM Mammalian Total RNA kit (Sigma) and treated with DNAse I according to the manufacturer's specifications. For each condition, three independent batches of RNA were prepared and controlled for purity and integrity using the Agilent 2100 Bioanalyzer with RNA 6000 Nano LabChip kits (Agilent Technologies). Only RNA with no sign of contamination or degradation (RIN > 9) were processed to generate amplified and biotinylated sense-strand cDNA targets using the GeneChip® WT PLUS Reagent kit from Affymetrix according to the manufacturer's specifications. After fragmentation, cDNA targets were used to probe Affymetrix GeneChip® Human Gene 2.0 ST arrays, which were then washed, stained and scanned according to Affymetrix instructions (manual P/N 702731 Rev.3).
5
Aortic gene expression analysis in OVX mice
6
RNA Isolation and Quality Control
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Total RNA was isolated using the miRNeasy Mini Kit (Qiagen) according to manufacturer's instructions. Aliquots of total RNA samples were used to determine the RNA concentration and purity using the Nano Drop ND-1000 spectral photometer (peqlab). RNA integrity was assessed by capillary electrophoresis using RNA 6000 Nano LabChip Kits and Bioanalyzer 2100 (Agilent Technologies). As recommended, total RNA samples with RNA integrity number (RIN) value ≥ 7.5 was used for the labelling the reaction to achieve high quality cRNA.
7
RNA Extraction, Labeling, and Analysis
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The RNA extraction, labeling, and analysis were performed by Shanghai Biotechnology Corporation. Total RNA was isolated using miRNeasy Mini Kit (Qiagen) according to the manufacturer’s protocol. RNA concentration, purity, and RNA integrity number (RIN) were measured using NanoDrop ND-1000 spectrophotometer (Peqlab, Erlangen, Germany), an Agilent 2100 Bioanalyzer and RNA 6000 Nano LabChip Kits (both Agilent Technologies, Santa Clara, CA, USA). A minimum RIN ≥ 6.0 was required for microarray analysis. Detailed methodology was from a previous publication [20 (link)].
8
Tumor Cell Enrichment for RNA Analysis
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The tumor cell content of a minimum of 80% of the biopsies was required, and manual microdissection was performed to enrich the tumor content when necessary. Total RNA, including small RNAs, was extracted using the FFPE miRNeasy Kit (Qiagen, Hilden, Germany). For each sample, between 2 and 10 unstained 10-µm sections were manually microdissected after deparaffinization, and RNA was extracted using 150 µl of proteinase K digestion buffer according to the manufacturer’s instructions. Total RNA was measured using the NanoDrop photospectrometer (NanoDrop, Wilmington, DE, USA) and was further processed if the A260/A280 ratio was ≥ 1.8 and the A260/A230 ratio was ≥ 1.4. All samples were analyzed using RNA 6000 Nano LabChip Kits (Agilent Technologies).
9
Transcriptomic Analysis of EYFP+ Cells
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RNA was isolated using TRIzol (Invitrogen) and RNeasy reagents (Qiagen). The integrity and concentration of RNA samples was determined using RNA 6000 Nano LabChip kits and an Agilent 2100 Bioanalyzer. Whole genome microarray analysis was performed on OneArray Mouse Whole Genome Array (Phalanx Biotech Group). The raw data were re-scaled to account for the differences in individual hybridization intensities. The heatmap represents relative expression as z-scores. The z-score of a gene was computed by using the expressions of all the genes both in EYFP+ and EYFP- cells. Gene Ontology analysis was performed on genes falling within the condition, p < 0.01, fold change >2, using DAVID (http://david.abcc.ncifcrf.gov ). GO network analysis was performed using enrichment Map (http://baderlab.org/Software/EnrichmentMap ) with p < 0.01 as the cutoff.
10
miR-9-3 Transcriptome Profiling in Mouse Cells
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RNA was extracted from mouse BMMCs and P815 cells transduced with empty lentivirus or pre-miR-9-3 lentivirus from three separate transduction experiments using TRIzol (Invitrogen). A secondary RNA cleanup step was performed using QIAGEN RNeasy Total RNA isolation kit (QIAGEN GmbH, Hilden, Germany) and RNA integrity was assessed using RNA 6000 Nano LabChip® Kits on the Agilent Bioanalyzer 2100 (Agilent Technologies, Palo Alto, CA, USA). RNA was labeled with Cy3 using RNA ligase and hybridized to GeneChip® Mouse Gene 2.0 ST Arrays (Affymetrix, Santa Clara, CA, USA). Ratios of signals were calculated and transcripts that were up-regulated or down-regulated by at least 2-fold were identified (p < 0.05). Data analysis, statistical analysis, and generation of gene expression heat maps were performed using Affymetrix® Transcriptome Analysis Console (TAC) Software. Prediction of miR-9 binding to the 3’-UTR of genes down-regulated by miR-9 was performed with computer-aided algorithms obtained from TargetScan (http://www.targetscan.org ), PicTar (http://pictar.mdc-berlin.de ), miRanda (http://www.microrna.org ), and miRWalk (http://www.umm.uni-heidelberg.de/apps/zmf/mirwalk ).
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