Rneasy kit

Manufactured by Promega

Sourced in United States

The RNeasy Kit is a laboratory equipment product designed for the purification of RNA from various biological samples. It utilizes a silica-membrane technology to efficiently capture and purify RNA molecules, making it suitable for a range of molecular biology applications.

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Lab products found in correlation

High capacity cdna reverse transcription kit, by Thermo Fisher Scientific (1 mentions) Steponeplus real time pcr system, by Thermo Fisher Scientific (1 mentions) Nanodrop photospectrometer, by Thermo Fisher Scientific (1 mentions) Dnase 1, by Promega (1 mentions) M mlv reverse transcriptase, by Promega (1 mentions) Sybr green, by Bio-Rad (1 mentions) Rneasy micro kit, by Qiagen (1 mentions) Chip seq sample prep kit, by Illumina (1 mentions) Ribo zero, by Illumina (1 mentions) Miniprep kit, by Qiagen (1 mentions) Reverse transcriptase kit, by Thermo Fisher Scientific (1 mentions) Lightcycler sybr green 1 master mix, by Roche (1 mentions) Dreamtaq polymerase, by Thermo Fisher Scientific (1 mentions) Verso cdna synthesis kit, by Thermo Fisher Scientific (1 mentions) Lightcycler 480, by Roche (1 mentions) Revertra ace kit, by Toyobo (1 mentions) Thunderbird sybr qpcr mix, by Toyobo (1 mentions) Cfx96, by Bio-Rad (1 mentions)

Close spelling variants of this product

RNeasy Mini Kit (26 citations) RNeasy Plant Mini Kit (6 citations) RNeasy extraction kit (3 citations) RNeasy plant kit (3 citations) RNeasy™ miniprep kit (2 citations) RNeasy Protect Bacteria mini kit (2 citations) RNeasy MinElute Cleanup Kit (2 citations)

9 protocols using rneasy kit

Quantitative gene expression analysis

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RNA was purified with the RNeasy Kit (Promega) and transcribed into cDNA (High Capacity cDNA Reverse Transcription Kit; Life Technologies). Real-time PCR was performed using the STEPOnePlus real-Time PCR system (Applied Biosystems). GAPDH and OAZ1 served as housekeeping genes for normalization. Primer sequences are listed in Table 1.

Yang W.J., Hu J., Uemura A., Tetzlaff F., Augustin H.G, & Fischer A. (2015). Semaphorin-3C signals through Neuropilin-1 and PlexinD1 receptors to inhibit pathological angiogenesis. EMBO Molecular Medicine, 7(10), 1267-1284.

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Quantifying Gene Expression in dgf Mutants

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Siblings were separated from dgf mutants at 7 dpf by using Alizarin live-staining. RNA was isolated by using the Qiagen RNeasy Kit, and a DNaseI (Promega) digest was performed on the column. RNA quality was checked on an agarose gel and measured with a Nanodrop photospectrometer (Thermo Scientific). Random hexamers were used for reverse transcription (M-MLV reverse transcriptase, Promega). The Primer 3 program was used for primer design. The primers spanned at least one intron to avoid amplification from genomic DNA. Melting temperatures and PCR efficiency were tested, and qPCR was performed using the Bio-Rad MyIQ single-color real-time PCR detection system and software. Reactions contained 12.5 μl Sybr Green (Bio-Rad), 3 μl primer mix (at 1.5 μM), 5 μl cDNA at 10 ng/μl, and 4.5 μl Millipore water. qPCR program: 3 minutes at 95°C, 10 seconds at 95°C followed by 45 seconds at the optimal primer temperature (40 cycles); 1 minute at 95°C and 1 minute at 65°C. cDNA was analyzed from three pooled clutches of embryos for the siblings and mutants. ef1a was used as an internal control. Groups were compared by Student’s t-test.

Apschner A., Huitema L.F., Ponsioen B., Peterson-Maduro J, & Schulte-Merker S. (2014). Zebrafish enpp1 mutants exhibit pathological mineralization, mimicking features of generalized arterial calcification of infancy (GACI) and pseudoxanthoma elasticum (PXE). Disease Models & Mechanisms, 7(7), 811-822.

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Total RNA Extraction and Gene Expression

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Total RNA was extracted using RNeasy micro Kit (Qiagen®) for rodent islets and RNeasy Kit (Promega®) for INS1 832/13 cells. cDNA was reversed transcribed. The levels of expression of each gene were normalized to cyclophylin expression (INS1832/13 and rat islets) and β-actin and cyclophylin (human islets).

Filhoulaud G., Benhamed F., Pagesy P., Bonner C., Fardini Y., Ilias A., Movassat J., Burnol A.F., Guilmeau S., Kerr-Conte J., Pattou F., Issad T, & Postic C. (2019). O-GlcNacylation Links TxNIP to Inflammasome Activation in Pancreatic β Cells. Frontiers in Endocrinology, 10, 291.

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ChIP-Seq and RNA-Seq in T47D Cells

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For chromatin immunoprecipitation-sequencing, a total of 5 × 10⁷ T47D cells were used per ChIP assay according to a previously described protocol^{35 (link)}. Briefly, cells were crosslinked with 1% paraformaldehyde for 10 min at room temperature, quenched with glycine, and fixed chromatin was sonicated and immunoprecipitated with specific antibodies. Libraries were prepared with Illumina’s ChIP-Seq sample prep kit for next-generation sequencing.
For RNA sequencing, T47D cells were transfected with control siRNA, TRPS1 siRNA, or CHD4 shRNA. Seventy-two hours later, total RNA was extracted with the RNeasy Kit (Promega). Total RNA was depleted of rRNA using Ribozero (Illumina), followed by library preparation and next-generation sequencing. The Gene Expression Omnibus (GEO) accession number for the ChIP-seq and expression data reported in this paper is GSE114213.

Wang Y., Lin X., Gong X., Wu L., Zhang J., Liu W., Li J, & Chen L. (2018). Atypical GATA transcription factor TRPS1 represses gene expression by recruiting CHD4/NuRD(MTA2) and suppresses cell migration and invasion by repressing TP63 expression. Oncogenesis, 7(12), 96.

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Cloning and Expressing PSAT1 in MCF-7 Cells

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Human PSAT1 was generated from cDNA prepared from the LCC9 cell line.
RNA was extracted using the RNeasy kit (Promega) and 1μg of RNA was used
to produce cDNA using the reverse transcriptase kit (Thermo) with random
primers. Primers homologous to the 5’ and 3’ ends of the mature
RNA encoding for PSAT1 were adapted to include restriction sites for ECOR1
(5’) and BAMH1 (3’). Polymerase chain reactions were performed to
amplify PSAT1 from LCC9 cDNA preparation. PCR products were separated via gel
electrophoresis and bands corresponding to the correct PCR product were excised
and purified. PSAT1 fragments underwent restriction digest with ECOR1 and BAMH1
and then ligated into the pcDNA3 vector that was similarly digested. Ligations
were then transformed and grown on LB agar plates with ampicillin. Isolated
colonies were cultured in LB broth and ampicillin for 24 hours and plasmids were
isolated via the Qiagen Miniprep kits. Plasmids, encoding either PSAT1 gene or
empty vector were verified by sequencing. Plasmids were then transfected into
the MCF-7 cell line using the Polyplus jetPRIME reagent according to
manufacturer’s protocol and clonal selection for PSAT1 expression was
performed using geneticin.

Metcalf S., Petri B.J., Kruer T., Green B., Dougherty S., Wittliff J.L., Klinge C.M, & Clem B.F. (2021). Serine Synthesis Influences Tamoxifen Response in ER+ Human Breast Carcinoma. Endocrine-related cancer, 28(1), 27-37.

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Semiquantitative RT-PCR Analysis of Oxidative Stress Genes

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Total RNA was extracted from kidney specimens by using the RNeasy kit (Promega, Madison, WI, USA) according to the manufacturer's instruction. The first strand cDNA was prepared using Superscript RT reverse transcriptase (Invitrogen, Carlsbad, CA). Semiquantitative RT-PCR assay of mRNA expression of HO-1, NQO1, Trx1and GAPDH genes was carried out as previously described [35 (link)] by using the following primers: HO-1, 5′-GGAACTTTCAGAAGGGCCAG-3' (forward) and 5′-GTCCTTGGTGTCATGGGTCA-3' (reverse); NQO1, 5′-CCCACAAGGTTGCAGCCGGA-3' (forward), 5′-CGGGCGTCTGCTGGAGTGTG-3' (reverse); Trx1, 5′-CGCCGGGCGTGCCAGTTTAT-3' (forward), 5′-TGGCTCCAGAAAATTCACCCACC-3' (reverse); GAPDH, 5′-CAATGCCTCCTGCACCACCA-3' (forward), 5′-GATGTTCTGGAGAGCCCCGC-3' (reverse). PCR amplification was conducted for a number of cycles in the linear range as determined in preliminary experiments. PCR products resolved in 1.5–2% agarose gels were photographed under ultraviolet light.

Lu M., Wang P., Qiao Y., Jiang C., Ge Y., Flickinger B., Malhotra D.K., Dworkin L.D., Liu Z, & Gong R. (2019). GSK3β-mediated Keap1-independent regulation of Nrf2 antioxidant response: A molecular rheostat of acute kidney injury to chronic kidney disease transition. Redox Biology, 26, 101275.

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Comprehensive RNA Analysis Workflow

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RNA was isolated (Qiagen RNeasy Kit) with subsequent DNase treatment (Promega) and reverse transcribed (Verso cDNA synthesis kit, Thermo Fisher). For qPCR, a LightCycler 480 from Roche was used with LightCycler SYBR Green I Master Mix. Semi-qPCR was performed using DreamTaq Polymerase (Thermo Fisher). For siRNA screen using MALT1 splicing-sensitive radioactive PCR, RNA was analysed with primers in flanking constant exons (ex6 and ex9/10). Products were analysed by denaturing PAGE and Phosphoimager quantification. PCR primers are listed in Supplementary Table 2 and 3.

Meininger I., Griesbach R.A., Hu D., Gehring T., Seeholzer T., Bertossi A., Kranich J., Oeckinghaus A., Eitelhuber A.C., Greczmiel U., Gewies A., Schmidt-Supprian M., Ruland J., Brocker T., Heissmeyer V., Heyd F, & Krappmann D. (2016). Alternative splicing of MALT1 controls signalling and activation of CD4+ T cells. Nature Communications, 7, 11292.

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Circadian Regulation of Brain RNA

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For RNA extraction, 8–10 weeks-old mice underwent a constant dark condition for 24–48 h. Brains were dissected, washed with HBSS, and immediately frozen with liquid nitrogen. RNA was extracted with RNeasy kit (Promega), and the cDNA library was created by first-strand reverse transcription with ReverTraAce Kit (Toyobo). For real-time quantitative PCR, cDNA samples were mixed with THUNDERBIRD SYBR qPCR Mix (Toyobo) and primers and underwent two-step PCR (95 ˚C 15 s→ 60 ˚C 60 s, 40 cycles).
Primer sequences for real-time quantitative PCR are explained in Supplementary Table 1.

Takeuchi S., Shimizu K., Fukada Y, & Emoto K. (2023). The circadian clock in the piriform cortex intrinsically tunes daily changes of odor-evoked neural activity. Communications Biology, 6, 332.

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RNA Extraction and qPCR Analysis

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RNA was isolated using a Qiagen RNeasy kit (Promega, Madison, WI). cDNA was prepared from 2 µg of total RNA following manufacturer's protocol. qPCR was performed using Taqman primers (Supplementary Table 1) using BioRad CFX96 for 40 cycles. The Ct value was determined by the ΔΔCT method normalized with β-actin. The sample numbers are shown in figure legends.

Chu F.F., Esworthy R.S., Doroshow J.H., Grasberger H., Donko A., Leto T.L., Gao Q, & Shen B. (2016). Deficiency in Duox2 activity alleviates ileitis in GPx1- and GPx2-knockout mice without affecting apoptosis incidence in the crypt epithelium. Redox Biology, 11, 144-156.

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