For extraction of total RNA or protein from OFC tissue samples, the miRVANA PARIS RNA extraction kit (Life Technologies, Carlsbad, CA) was used following the manufacturer’s instructions, as previously reported [64 (link)]. OFC was collected from the ventral (VO) and lateral subregions (LO). The RNA fraction was suspended in RNase free water and the RNA concentration was measured with a Qubit 3.0 Fluorometer and the Qubit RNA High Sensitivity Assay (Invitrogen, Carlsbad, California, USA). Protein fractions were suspended in 1X RIPA buffer (5X RIPA buffer: 100 mM Tris-HCl, 750 mM NaCl, 5 mM ethylenediaminetetraacetic acid [EDTA], 5 mM ethylene glycol-bis[β-aminoethyl ether]-N,N,N’,N’-tetraacetic acid [EGTA], 5% sodium deoxycholate, 0.5% sodium dodecyl sulfate [SDS]) and 1X Protease and Phosphatase Inhibitor Cocktail (Thermo Scientific).
Mirvana paris rna extraction kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The MiRVANA PARIS RNA extraction kit is a tool used for the isolation and purification of total RNA from a variety of sample types, including cells, tissues, and body fluids. The kit utilizes a proprietary method to efficiently extract high-quality RNA for downstream applications.
Automatically generated - may contain errors
Lab products found in correlation
Norgen rna dna protein purification plus kit, by Norgen Biotek (2 mentions)
Protease and phosphatase inhibitor cocktail, by Thermo Fisher Scientific (1 mentions)
Qubit 3.0 fluorometer, by Thermo Fisher Scientific (1 mentions)
Qubit rna high sensitivity assay, by Thermo Fisher Scientific (1 mentions)
Mircury lna sybr green pcr kit, by Qiagen (1 mentions)
Lna sybr green primers, by Qiagen (1 mentions)
Mircury lna rt kit, by Qiagen (1 mentions)
5 protocols using mirvana paris rna extraction kit
1
RNA and Protein Extraction from OFC Tissue
2
RNA Extraction for Rodent and Human Tissues
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For rodent studies, total RNA was extracted from fresh frozen bilateral tissue punches using the miRVANA PARIS RNA extraction kit (Life Technologies, Carlsbad, CA). For human studies, total RNA was extracted using the Norgen RNA/DNA/Protein Purification Plus Kit (Norgen Biotek, Ontario, Canada) according to the manufacturer’s protocol. For additional details, please see supplemental methods .
3
RNA Extraction for Rodent and Human Tissues
4
RNA Extraction with miRVANA PARIS Kit
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Total RNA, including small RNAs was extracted using the miRVANA PARIS RNA extraction kit (Life Technologies, CA, USA), according to the manufacturer’s protocol. For plasma samples, each sample was spiked with 5 μL of 200 nM synthetic cel-miR-39 and ath-miR159a (Sigma-Aldrich, MO, USA) after RNases were inactivated. The extraction protocol was modified with an extended centrifugation time of 60 min at 13,000×g at the organic extraction step, and each RNA sample was eluted in 50 μL of hot elution (95 °C) buffer.
5
Quantification of miRNA Expression
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Total RNA from fresh frozen bilateral tissue punches was obtained using the miRVANA PARIS RNA extraction kit (Life Technologies), as previously reported (Rumbaugh et al. 2015 (link); Sillivan et al. 2019b (link)). cDNA libraries of miRNAs were created from 20 ng of total RNA with the mirCURY LNA RT Kit (Qiagen). PCR reactions were performed using the miRCURY LNA SYBR Green PCR Kit and the following locked nucleic acid (LNA) SYBR green primers from Qiagen: mmu-miR-598-3p, assay ID: YP00205045; snord68, assay ID: YP00203911; and RNU5G, assay ID: YP00203908. Data were normalized to the housekeeping genes snord68 and RNU5G using the ΔΔCt method (Livak and Schmittgen 2001 (link)).
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