Cy3-labeled cDNA was prepared from total RNA, according to the protocol described in the NimbleGen Arrays User’s Guide version 5.0 (Roche NimbleGen Inc., Madison, WI, USA). All samples were prepared in the same manner. Cy3-labeled cRNA was quantified in a spectrophotometer (Amersham Pharmacia Biotech, GE Healthcare Bio-Sciences AB, Uppsala, Sweden) and was fragmented following the manufacturer recommendations. Fragmented cDNA was hybridized to the NimbleGen A. oryzae custom-designed array following the manufacturer recommendations. Arrays were washed and stained using a GeneChip® Fluidics Station FS-400, and scanned on an Agilent GeneArray® Scanner 3000. The scanned probe array images were converted into data files using the NimbleScan software.
Nimblegen array
Manufactured by Roche
Sourced in United States, Switzerland
NimbleGen Arrays are a high-density microarray platform designed for genome-wide analysis. They provide a comprehensive solution for DNA and RNA profiling, including gene expression, copy number variation, and chromatin immunoprecipitation (ChIP) studies. The arrays feature a flexible design and high-density probe coverage to enable detailed, accurate, and reproducible genomic analysis.
Automatically generated - may contain errors
Lab products found in correlation
Superscript double stranded cdna synthesis kit, by Thermo Fisher Scientific (2 mentions)
Nimblegen one color dna labeling kit, by Thermo Fisher Scientific (2 mentions)
Agilent microarray, by Agilent Technologies (1 mentions)
Agilent kit, by Agilent Technologies (1 mentions)
Genepix 4000b scanner, by Molecular Devices (1 mentions)
Signalmap v 1, by Roche (1 mentions)
Total rna isolation kit, by Promega (1 mentions)
Dnase kit, by Thermo Fisher Scientific (1 mentions)
Transcriptor high fidelity cdna synthesis kit, by Roche (1 mentions)
Agilent bioanalyzer, by Agilent Technologies (1 mentions)
Sigma wta2 whole transcriptome amplification kit, by Merck Group (1 mentions)
Nimblescan software, by Roche (1 mentions)
6 protocols using nimblegen array
1
Transcriptome Analysis of Aspergillus oryzae
2
Comparative Microarray Analysis of Tumor DNA
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For each sample having sufficient quality and quantity, 700 to 1000 ng of tumor DNA and reference DNA were labeled, purified, and co-hybridized in equal quantity either to the NimbleGen Arrays (Roche, Basel, Switzerland) or Agilent Microarrays (Agilent Technologies, Santa Clara, CA, USA), during 12 to 24 h. Male and female human reference DNA were extracted from human blood for those analyzed on a NimbleGen support, whereas references were provided in the Agilent Kit for those analyzed on an Agilent support. Arrays were washed and scanned according to the technique-specific guidelines.
For samples processed with the Nimblegen technology, images were acquired on a GenePix 4000B scanner with the GenePix V.6.6 software (Molecular Devices, San Jose, CA, USA), and data was extracted using the NimbleScan V.2.5 software. Files produced by the NimbleScan software were then analyzed on SignalMap V.1.9 (Roche, Basel, Switzerland).
For samples processed with the Agilent technology, images were acquired on a SureScan Microarray Scanner using CytoScan software V.2.7, and then analyzed on CytoGenomics software V.3.0.2.11 (Thermo Fisher Scientific, Waltham, MA, USA).
For each sample, the quality of the analysis was evaluated subjectively based on the cytogenetic profile dynamism, the sex mismatch, and the degree of dispersion.
For samples processed with the Nimblegen technology, images were acquired on a GenePix 4000B scanner with the GenePix V.6.6 software (Molecular Devices, San Jose, CA, USA), and data was extracted using the NimbleScan V.2.5 software. Files produced by the NimbleScan software were then analyzed on SignalMap V.1.9 (Roche, Basel, Switzerland).
For samples processed with the Agilent technology, images were acquired on a SureScan Microarray Scanner using CytoScan software V.2.7, and then analyzed on CytoGenomics software V.3.0.2.11 (Thermo Fisher Scientific, Waltham, MA, USA).
For each sample, the quality of the analysis was evaluated subjectively based on the cytogenetic profile dynamism, the sex mismatch, and the degree of dispersion.
3
Transcriptomic Response of P. gingivalis to DEA NONOate
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As in NO sensitivity experiment, culture samples were taken 15 min posttreatment with DEA NONOate from growing P. gingivalis W83 cultures and total RNA was extracted from them using a total RNA isolation kit (Promega, Madison, WI). Additional DNase treatment was carried out using DNase kit (Thermo Fisher Scientific, Waltham, MA). Samples from untreated cultures of P. gingivalis W83 were processed similarly and used as controls. DNA microarray gene expression was carried out using Roche NimbleGen customer arrays (100910_CW_P_ging_W83_expr_HX12; Roche Diagnostics, Indianapolis, IN) according to the standard NimbleGen procedure (NimbleGen Arrays User’s Guide: Gene Expression Analysis v5.1). cDNA was synthesized from the RNA samples using Transcriptor High Fidelity cDNA Synthesis kit (Roche Diagnostics, Indianapolis, IN). Both RNA and cDNA qualities were checked using Agilent Bioanalyzer and Agilent RNA 6000 Nano and DNA1000 chips. Microarray procedures were carried out as previously outlined (Boutrin et al., 2012 (link)). Differentially expressed genes (DEGs) were determined using fold change (≥1.25) plus p (≤0.05) with a false discovery rate (FDR) of 0.05. The microarray data were submitted to NCBI’sGene Expression Omnibus database and are accessible through GEO Series accession number GSE224067 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE224067 ).
4
Genome-wide Expression Analysis of miR-204
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Global gene expression analysis was done for MDA-MB-231 cells transfected with pre-miR-204 (30 nM) or scramble (30 nM) using the NimbleGen array (Roche). RNA samples were used to synthesize double-stranded labeled cDNA using SuperScript Double-Stranded cDNA Synthesis Kit (Invitrogen) and NimbleGen One-Color DNA Labeling Kit. Samples were hybridized in NimbleGen array 12 × 135 K (12 × 135,000 features). After hybridization and washing, the processed slides were scanned using a NimbleGen MS200 Microarray Scanner. Raw data were extracted as pair files by NimbleScan software (version 2.5), background corrected and data were normalized. The probe level files and gene summary files were produced and imported into ANAIS software (Analysis of NimbleGen arrays Interface) for further analysis. DNA microarrays data for 10 selected genes was validated by RT-qPCR using specific oligonucleotides.
5
Genome-wide Analysis of miR-944 Regulation
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Global gene expression analysis was done for MDA-MB-231 cells transfected with miR-944 precursor (50 nM) or scramble (30 nM) using the NimbleGen array (Roche). RNA samples were used to synthesize double-stranded labeled cDNA using SuperScript Double-Stranded cDNA Synthesis Kit (Invitrogen) and NimbleGen One-Color DNA Labeling Kit. Samples were hybridized in NimbleGen array 12x135K (12 x 135,000 features). After hybridization and washing, the processed slides were scanned using a NimbleGen MS200 Microarray Scanner. Raw data were extracted as pair files by NimbleScan software (version 2.5), background was corrected and data were normalized. The probe level files and gene summary files were produced and imported into ANAIS software (Analysis of NimbleGen arrays Interface) for further analysis. The Student test with Varmixt package was used and raw P values were adjusted by the Benjamini and Yekutieli method to control the false discovery rate (FDR). Only genes with a Benjamini/Yekutieli value <0.05, and expression fold change >1.5 were considered as being differentially expressed.
6
Whole Transcriptome Amplification and Microarray Analysis
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300ηg of RNA was used as starting material for cDNA synthesis using the Sigma WTA2 whole transcriptome amplification kit as previously described [17 (link)].
1μg of cDNA was labeled with Cy3 dye using 65% AT rich pre-labeled random hexamers as primers for cDNA synthesis by Klenow fragment of DNA polymerase I. Hybridizations on the Nimblegen array were performed for 18 hours followed by washing of the arrays as described according to standard protocol (Roche NimbleGen Inc., Madison, WI). The microarray image was obtained using a 2μM scanner and probe intensity values extracted using NimbleScan software (Roche NimbleGen Inc., Madison, WI). For the Agilent HD exon array, hybridizations were performed for 17 hours. Images were obtained on the same 2uM scanner and probe intensity values extracted using Agilent Feature Extraction software.
1μg of cDNA was labeled with Cy3 dye using 65% AT rich pre-labeled random hexamers as primers for cDNA synthesis by Klenow fragment of DNA polymerase I. Hybridizations on the Nimblegen array were performed for 18 hours followed by washing of the arrays as described according to standard protocol (Roche NimbleGen Inc., Madison, WI). The microarray image was obtained using a 2μM scanner and probe intensity values extracted using NimbleScan software (Roche NimbleGen Inc., Madison, WI). For the Agilent HD exon array, hybridizations were performed for 17 hours. Images were obtained on the same 2uM scanner and probe intensity values extracted using Agilent Feature Extraction software.
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