Ovation pico wta systems v2

Manufactured by Tecan

The Ovation® Pico WTA Systems V2 is a laboratory equipment designed for whole transcriptome amplification from low-input RNA samples. It provides a streamlined workflow for generating amplified cDNA from as little as 500 picograms of total RNA.

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Lab products found in correlation

Encore biotinil module, by Tecan (2 mentions) Rna 6000 nano chip, by Agilent Technologies (1 mentions) Human genome u133 plus2 gene chip, by Thermo Fisher Scientific (1 mentions) 3 ivt express kit, by Thermo Fisher Scientific (1 mentions) Trizol, by Thermo Fisher Scientific (1 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions) Iscan array scanner, by Illumina (1 mentions) Easysep positive selection kit, by STEMCELL (1 mentions) Ingenuity pathway analysis ipa, by Qiagen (1 mentions) Genomestudio, by Illumina (1 mentions) Rneasy mini kit, by Qiagen (1 mentions)

2 protocols using ovation pico wta systems v2

Transcriptomic profiling of primary cells and cell lines

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For primary cell experiments, cell pellets for mRNA isolation were lysed in Trizol (Ambion, Life Technologies) and RNA extracted by guanidium-isothiocyanate phenol/chloroform extraction method following the manufacturer’s instructions. Total RNA was quantified using the Nanodrop ND1000 and RNA integrity assessed with the Agilent Bioanalyzer 2100 using the RNA 6000 Nano Chips. One hundred nanograms of total RNA was used to prepare the targets, using the 3′ IVT Express kit (Affymetrix) in accordance with the manufacturer’s instructions. Hybridisation cocktails were hybridised onto the Human Genome U133 plus2 gene chip (Affymetrix). For Jurkat cell line experiments, three technical replicates of Jurkat T cell stably expressing CYT-1 or CYT-2 and untransduced cells as control were analysed. RNA was converted to cDNA using the Ovation® Pico WTA Systems V2 and labelled using the Encore® BiotinIL Module (NuGen) according to the manufacturers recommendations. Labelled cDNA was hybridised onto Illumina HT12v4 arrays (Illumina).

Perucha E., Melchiotti R., Bibby J.A., Wu W., Frederiksen K.S., Roberts C.A., Hall Z., LeFriec G., Robertson K.A., Lavender P., Gerwien J.G., Taams L.S., Griffin J.L., de Rinaldis E., van Baarsen L.G., Kemper C., Ghazal P, & Cope A.P. (2019). The cholesterol biosynthesis pathway regulates IL-10 expression in human Th1 cells. Nature Communications, 10, 498.

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Transcriptomic Profiling of CD19+ B Cells in Sepsis

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CD19+ B cells from sepsis and healthy controls were isolated to a purity of >99% using EasySep ® Positive Selection kit (Stem Cell Technologies). Purity was confirmed with flow cytometry. Total RNA was isolated using the RNeasy Mini Kit (Qiagen). RNA was converted to cDNA using the Ovation ® Pico WTA Systems V2 and labelled using the Encore ® BiotinIL Module (Nugen). Labelled cDNA was hybridized onto Infinium Illumina HT12v4 arrays and data collected on an iSCAN array scanner (Illumina). Following scanning, basic QC statistics were completed, displaying hybridisation controls, background signal and mean gene intensity of all genes and those of housekeeping genes. Array results were compiled using Genome Studio (Illumina) following quantile normalisation. Differences in messenger RNA abundance between the sepsis and healthy controls were assessed using Partek Genomic Suite (Partek Inc.). Similarities and differences in cell specific gene expression pattern in health and sepsis were assessed using heat maps and the top canonical pathways identified using Ingenuity Pathway Analysis (IPA) (Qiagen). Apoptosis genes and pathways were also studied.

Shankar-Hari M., Fear D., Lavender P., Mare T., Beale R., Swanson C., Singer M, & Spencer J. (2017). Activation-Associated Accelerated Apoptosis of Memory B Cells in Critically Ill Patients With Sepsis. Critical care medicine, 45(5).

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