Total RNA was purified with GenElute total RNA kit (Sigma-Aldrich). RNA from PS-like structures and SG cores was purified using TRI-reagent (Sigma-Aldrich). First-strand cDNA synthesis was performed using random primers (Promega) and Superscript IV (Invitrogen) according to the manufacturers’ instructions. PCR was performed using New England BioLabs Taq DNA polymerase (M0273). Quantitative real-time PCR was run in triplicate on a StepOne real-time PCR instrument, and data were analyzed using StepOne software v2.0 (Applied Biosystems). GAPDH was used for normalization. Primer sequences were as follows: NEAT1 total, 5′-CTCACAGGCAGGGGAAATGT-3′ and 5′-AACACCCACACCCCAAACAA-3′; NEAT1_2, 5′-TGTGTGTGTAAAAGAGAGAAGTTGTGG-3′ and 5′-AGAGGCTCAGAGAGGACTGTAACCTG-3′; GAPDH, 5′-TCGCCAGCCGAGCCA-3′ and 5′-GAGTTAAAAGCAGCCCTGGTG-3′.
Genelute total rna kit
Manufactured by Merck Group
The GenElute total RNA kit is a laboratory product designed for the extraction and purification of total RNA from various biological samples. It provides a reliable and efficient method for isolating high-quality RNA for downstream applications such as RT-PCR, Northern blotting, and gene expression analysis.
Automatically generated - may contain errors
Lab products found in correlation
Lightcycler 480, by Roche (2 mentions)
Taqman master mix, by Thermo Fisher Scientific (2 mentions)
Cdna transcription kit, by Thermo Fisher Scientific (2 mentions)
Tri reagent, by Merck Group (1 mentions)
Stepone software v2, by Thermo Fisher Scientific (1 mentions)
Taq dna polymerase, by New England Biolabs (1 mentions)
Real time pcr instrument, by Thermo Fisher Scientific (1 mentions)
Random primer, by Promega (1 mentions)
Superscript 4, by Thermo Fisher Scientific (1 mentions)
3 protocols using genelute total rna kit
1
NEAT1 isoforms expression analysis
2
Quantitative Analysis of Human OPC and Microglia
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Human OPC cDNA extracted from normal primary cells was purchased from ScienCell (1604, Carslbad, CA, USA). Human microglia cell total RNA extracted from primary cells was purchased from Celprogen (37089RNA, Torrance, CA, USA). Treated MO3.13 oligodendrocytes were washed in PBS, scraped, spun for 10 min at maximum speed and resuspended in 150 μL of lysis buffer (Sigma-Aldrich, RTN70-1KT). Total RNA was extracted using the GenElute total RNA kit (Sigma-Aldrich, RTN70-1KT). Isolated total RNA was frozen at −20 °C until used. Total RNA was reverse-transcribed with cDNA transcription kit (ThermoFisher, 4368814) according to the manufacturer’s instructions. RT-qPCR was performed with TaqMan master mix (ThermoFisher, 4444556) using the LightCycler 480 (Roche, Basel, Switzerland) according to the standard protocol. FAM dye-labelled TaqMan (Applied Biosystems, Foster City, CA, USA) probes were used in all experiments. The relative mRNA expression using the ΔΔCt method was calculated from absolute quantification after normalization to the reference gene.
3
Quantitative RT-qPCR in Cuprizone Mouse Model
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The snap-frozen mouse hemispheres from the cuprizone model were homogenised in lysis solution (Sigma-Aldrich, L8285) and total RNA was extracted using the GenElute total RNA kit (Sigma-Aldrich, RTN70-1KT) according to the manufacturer's instruction. The concentration of RNA was quantified, equalised and reverse transcribed with cDNA transcription kit (ThermoFisher, 4368814) according to the manufacturer's instructions. RT-qPCR was performed using TaqMan master mix (ThermoFisher, 4444556) on the LightCycler480 (Roche) according to the standard protocol. FAM dye-labelled TaqMan (AppliedBiosystems, 4331182) probes were used in all experiments. The relative mRNA expression using the ΔΔCt method was calculated from absolute quantification after normalization to the reference gene.
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