Magattract powermicrobiome dna rna ep kit

Manufactured by Qiagen

Sourced in United States

The MagAttract PowerMicrobiome DNA/RNA EP Kit is a laboratory equipment product designed for the extraction and purification of microbial DNA and RNA from a variety of sample types. The kit utilizes magnetic bead technology to efficiently capture and separate the nucleic acids from the sample matrix.

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Lab products found in correlation

Nextera dna flex library prep kit, by Illumina (4 mentions) Agilent 4200 tapestation, by Agilent Technologies (3 mentions) Epmotion 5073, by Eppendorf (3 mentions) Nextseq 500 instrument, by Illumina (2 mentions) Dsdna high sensitivity assay, by Agilent Technologies (2 mentions) Agilent high sensitivity hs d5000 screentape assay, by Agilent Technologies (2 mentions) Qubit 4 fluorometer, by Thermo Fisher Scientific (2 mentions) Agilent high sensitivity d5000 screentape, by Agilent Technologies (1 mentions) Qubit dsdna hs assay, by Thermo Fisher Scientific (1 mentions) Nextseq 500 sequencer, by Illumina (1 mentions) Synergy htx plate reader, by Agilent Technologies (1 mentions) Lysis matrix e tubes, by MP Biomedicals (1 mentions) Qubit 2.0 fluorometer, by Thermo Fisher Scientific (1 mentions) Precellys 24 tissue homogenizer, by Bertin Technologies (1 mentions) Nextseq 500, by Illumina (1 mentions) Kingfisher duo, by Thermo Fisher Scientific (1 mentions) Accuprime taq dna polymerase, by Thermo Fisher Scientific (1 mentions) Sequalprep normalization plate kit, by Thermo Fisher Scientific (1 mentions) Quant it picogreen dsdna assay kit, by Thermo Fisher Scientific (1 mentions) Miseq system, by Illumina (1 mentions)

8 protocols using magattract powermicrobiome dna rna ep kit

Metagenomic Analysis of Stool Samples

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Stool samples were collected at different time points per individual (Supplementary Table 3). DNA was extracted from ~50 mg of stool samples in two stages: an initial homogenization in Lysis Matrix E tubes (MP Biomedicals) with a Precellys 24 Tissue Homogenizer (Bertin Instruments) and processing of the resultant supernatant using the MagAttract PowerMicrobiome DNA/RNA EP kit (Qiagen) on an Eppendorf automated liquid handling system as per the manufacturer’s instructions.
Isolated DNA was checked for concentration and quality on a BioTek Synergy HTX plate reader.
Metagenomic libraries were prepared using the Nextera DNA Flex Library Prep Kit (Illumina) per the manufacturer’s instructions with 100 ng of DNA as sample input. The concentration of the libraries was quantified using the Qubit dsDNA HS assay on a Qubit 2.0 fluorometer (Life Technologies). Library size and quality were assessed via the Agilent High Sensitivity D5000 ScreenTape on an Agilent 4200 Tapestation.
Metagenomic libraries were normalized to an equimolar concentration and pooled. The pool was diluted to 1.8 pM, mixed with a 1% PhiX control library and paired-end sequenced (2 × 75 bp) using a NextSeq 500/550 High Output v2 150-cycle Reagent Cartridge on a NextSeq 500 sequencer (Illumina).

Link V.M., Subramanian P., Cheung F., Han K.L., Stacy A., Chi L., Sellers B.A., Koroleva G., Courville A.B., Mistry S., Burns A., Apps R., Hall K.D, & Belkaid Y. (2024). Differential peripheral immune signatures elicited by vegan versus ketogenic diets in humans. Nature Medicine, 30(2), 560-572.

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Metagenomic DNA Extraction and Sequencing

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DNA was isolated using the MagAttract PowerMicrobiome DNA/RNA EP Kit (Qiagen) in an Eppendorf epMotion 5073 automated liquid handling system. Metagenomic libraries were prepared using 30 μl DNA in a 96-well plate with the Nextera DNA Flex Library Prep Kit (Illumina). The library preparation was initiated with tagmentation and post-tagmentation cleanup followed by amplification of tagmented DNA and cleanup using Sample Purification Beads for both the cleanup steps. Library quality control included quantification of the final library using the Qubit dsDNA High Sensitivity assay and library quality assessment using the Agilent High Sensitivity HS D5000 ScreenTape assay on the Agilent 4200 Tapestation. The individual libraries were diluted and pooled at a concentration of 7 nM, to make the final library pool. This pool was normalized to 1.8 pM, spiked-in with 1% PhiX sequencing control and sequenced on an Illumina NextSeq 500 instrument.

Ozen A., Kasap N., Vujkovic-Cvijin I., Apps R., Cheung F., Karakoc-Aydiner E., Akkelle B., Sari S., Tutar E., Ozcay F., Uygun D.K., İslek A., Akgun G., Selcuk M., Sezer O.B., Zhang Y., Kutluk G., Topal E., Sayar E., Celikel Ç., Houwen R.H., Bingol A., Ogulur I., Eltan S.B., Snow A.L., Lake C., Fantoni G., Alba C., Sellers B., Chauvin S.D., Dalgard C.L., Harari O., Ni Y.G., Wang M.D., Devalaraja-Narashimha K., Subramanian P., Ergelen R., Artan R., Guner S.N., Dalgic B., Tsang J., Belkaid Y., Ertem D., Baris S, & Lenardo M.J. (2021). Broadly effective metabolic and immune recovery with C5 inhibition in CHAPLE disease. Nature immunology, 22(2), 128-139.

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Metagenomic DNA Extraction and Sequencing

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Fecal Metagenomics: Microbiome Analysis

Cited 3 times

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Fecal microbiome was analyzed as previously described^{53 (link)}. DNA was isolated using the MagAttract PowerMicrobiome DNA/RNA EP Kit (Qiagen) in an Eppendorf epMotion 5073 automated liquid handling system. Metagenomic libraries were prepared using 30 μl of DNA with the Nextera DNA Flex Library Prep Kit (Illumina). Library preparation was initiated with tagmentation and post-tagmentation cleanup followed by amplification of tagmented DNA and cleanup using Sample Purification Beads for both cleanup steps. The individual libraries were diluted and pooled at a concentration of 7 nM to make the final library. This pool was normalized to 1.8 pM, spiked-in with 1% PhiX sequencing control and sequenced on an Illumina NextSeq 500 instrument. For sequence analysis, MetaPhlAn2 was used to obtain relative abundances. Principal coordinates analysis plots were generated using the Canberra distance metric based on resulting species-level classifications^{54 (link)}. For estimating percentages of Human Microbiome Project (HMP) body site origin, the SourceTracker^{55 (link)} approach was applied by training on MetaPhlAn data from the HMP. Alpha values were tuned via cross-validation, and SourceTracker analyses were performed on genus-level taxonomic classifications.

Kubo S., Fritz J.M., Raquer-McKay H.M., Kataria R., Vujkovic-Cvijin I., Al-Shaibi A., Yao Y., Zheng L., Zou J., Waldman A.D., Jing X., Farley T.K., Park A.Y., Oler A.J., Charles A.K., Makhlouf M., AbouMoussa E.H., Hasnah R., Saraiva L.R., Ganesan S., Al-Subaiey A.A., Matthews H., Flano E., Lee H.H., Freeman A.F., Sefer A.P., Sayar E., Çakır E., Karakoc-Aydiner E., Baris S., Belkaid Y., Ozen A., Lo B, & Lenardo M.J. (2021). Congenital iRHOM2 deficiency causes ADAM17 dysfunction and environmentally directed immunodysregulatory disease. Nature immunology, 23(1), 75-85.

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Duodenal Aspirate and Stool DNA Isolation

Cited 1 time

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On the day of DNA isolation, duodenal aspirate microbial pellets under Allprotect were thawed on ice and 1x DTT was added in a 1:1 ratio and the mixture was vortexed.³¹ (link) Microbial DNAs were isolated from duodenal aspirates using the MagAttract PowerMicrobiome DNA/RNA EP Kit (Qiagen) on a KingFisher Duo (Thermo Fisher Scientific, Waltham, MA, USA).³¹ (link) Stool DNAs were isolated using MagAttract PowerSoil DNA EP Kits (Qiagen) and purified using a KingFisher Duo automated system (ThermoFisher Scientific, Waltham, MA). Isolated DNAs were quantified using Qubit 1X dsDNA, High Sensitivity Assay kits (Invitrogen by Thermo Fisher Scientific) on a Qubit 4 Fluorometer (Invitrogen, Carlsbad, CA, USA).

Hosseini A., Barlow G.M., Leite G., Rashid M., Parodi G., Wang J., Morales W., Weitsman S., Rezaie A., Pimentel M, & Mathur R. (2023). Consuming artificial sweeteners may alter the structure and function of duodenal microbial communities. iScience, 26(12), 108530.

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Fecal Microbiome Profiling via 16S rRNA

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DNA was extracted from feces pellets using the Qiagen MagAttract PowerMicrobiome DNA/RNA EP kit (QIAGEN, catalog 27500-4-EP). The V4 region of the 16S rRNA-encoding gene was amplified from extracted DNA using the barcoded dual-index primers developed by Kozich et al. (61 (link)) (Supplemental Table 8). Samples were amplified using AccuPrime Taq DNA Polymerase, high fidelity (Invitrogen, catalog 12346086) at 95°C for 2 minutes followed by 30 cycles of 95°C for 20 seconds, 55°C for 15 seconds, and 72°C for 5 minutes, and final incubation at 72°C for 10 minutes, purified using a magnetic bead capture kit (Agencourt AMPure; Beckman Coulter, catalog 000130) and quantified using a fluorometric kit (Quant-iT PicoGreen dsDNA Assay Kit; Invitrogen, catalog P7589). Purified amplicons were pooled in equimolar concentrations with a SequalPrep Normalization Plate Kit (Applied Biosystems, catalog A1051001) and sequenced on Illumina MiSeq System (RRID:SCR_016379). Bioinformatic analysis was done using the Mothur v.1.42.3 software package (62 (link)) (RRID:SCR_011947) available at the University of Michigan Microbial Systems Laboratory.

Platt J.L., de Mattos Barbosa M.G., Huynh D., Lefferts A.R., Katta J., Kharas C., Freddolino P., Bassis C.M., Wobus C., Geha R., Bram R., Nunez G., Kamada N, & Cascalho M. (2021). TNFRSF13B polymorphisms counter microbial adaptation to enteric IgA. JCI Insight, 6(14), e148208.

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Soil Microbiome DNA/RNA Extraction and 16S Sequencing

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Plant root tissues in the soil samples were removed to minimize plant nucleic acid contamination. All soil samples were homogenized prior to DNA and RNA extraction and subsampled to 0.25 g. DNA and RNA extraction was performed using the MagAttract PowerMicrobiome DNA/RNA EP Kit (Qiagen, USA) following the standard protocol in this kit and liquid handling in Eppendorf epMotion 5075 (Eppendorf North America). The extracted RNA was transcribed into cDNA according to a standard protocol using iScript cDNA Synthesis Kit (BIO-RAD, USA) for sequencing analysis. The resulting DNA and RNA were analyzed for quantity using an Invitrogen Qubit 4 Fluorometer (Invitrogen, USA). DNA and RNA sample concentrations above 10 ng μL⁻¹ were normalized to 10 ng μL⁻¹ prior to sequencing. Samples with concentrations lower than 10 ng μL⁻¹ were submitted directly for sequencing. The V4 region of the bacterial 16S rRNA gene was amplified with the conserved primers 515F (5′-GTGYCAGCMGCCGCGGTAA-3′) and 806R (5′-GGACTACNVGGGTWTCTAAT-3′) (76 (link), 77 (link)). Bacterial amplicon sequencing was performed on Illumina Miseq with Miseq reagent kit V2 (Illumina, USA) at Argonne National Laboratory.

Yang J., Lee J., Choi J., Ma L., Heaton E.A, & Howe A. (2022). Response of Total (DNA) and Metabolically Active (RNA) Microbial Communities in Miscanthus × Giganteus Cultivated Soil to Different Nitrogen Fertilization Rates. Microbiology Spectrum, 10(1), e02116-21.

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Microbiome DNA Extraction and Quantification

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Piglet and sow faecal samples, mock community samples, negative controls and probiotic samples (D-Scour™ and ColiGuard® paste) were allocated to a randomized block design to control for batch effects in DNA extraction and library preparation. The faecal samples were thawed on ice rst, followed by the probiotics and mock community samples. MetaPolyzyme (Sigma-Aldrich) treatment was performed according to the manufacturer's instructions except for the dilution factor, which we allowed to be 4.6 times higher. Immediately after incubation, DNA extraction was performed with the MagAttract PowerMicrobiome DNA/RNA EP kit (Qiagen) according to the manufacturer's instructions. Quanti cation of DNA was performed using PicoGreen (Thermo sher) and measurements were performed with a plate reader (Tecan, Life Sciences) using 50 and 80 gain settings. All samples were diluted to 10 ng/µL.

Gaio D., DeMaere M.Z., Anantanawat K., Eamens G.J., Liu M., Zingali T., Falconer L., Chapman T.A., Djordjevic S.P., & Darling A.E. (2020). Community composition and development of the post-weaning piglet gut microbiome.

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