Monarch® Mag Viral DNA/RNA Extraction Kit |NEB酶试剂 New England Biolabs

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产品信息

The Monarch Mag Viral DNA/RNA Extraction Kit provides a rapid and reliable magnetic bead-based process for extracting viral nucleic acids from saliva and respiratory swab samples. The kit combines the efficiency of silica-based nucleic acid purification with the ease of use of magnetic beads. Manual and automated workflows allow samples to be processed in microfuge tubes or 96-well plates. Kit sizes align to 96-well formats (100 preps, 600 preps, and 1800 preps), and the protocol is compatible with high throughput automation on a variety of platforms, including the KingFisher Flex magnetic particle processor and Agilent Bravo and MGISP liquid handler platforms.

Properties
Purification format Magnetic bead
Processing format Manual or automated
Sample purification (representative examples) Viral DNA and RNA* from respiratory viruses (enveloped and non-enveloped, dsDNA and ssRNA)
Sample sources Saliva, respiratory swab in viral transport media (VTM)**
Sample input volume Up to 200 μl**
Carrier supplier Poly A carrier RNA***
Binding capacity Up to 3 μg
Elution volume 33–100 μl
Tested automation platforms KingFisher Flex; Agilent Bravo and MGISP liquid handlers
Compatible downstream applications qPCR, RT-qPCR, ddPCR, library prep for NGS

* Viral DNA and RNA are purified in parallel. Preparation of DNA-free RNA or RNA-free DNA requires further treatment with the appropriate nuclease (not supplied).
** The sample input volume may be scalable to accommodate larger sample volumes. Further workflow optimization may be required.
*** Use of carrier RNA is recommended for recovery of low amounts of viral nucleic acid. Carrier RNA should be omitted if the downstream application utilizes poly(A) RNA enrichment; however, viral nucleic acid recovery may be reduced.

 

Figure 1: Monarch Mag Viral DNA/RNA Extraction Kit Workflow

Monarch® Mag Viral DNA/RNA Extraction Kit |
Overview of workflow of the Monarch Mag Viral DNA/RNA Extraction Kit for manual or automated magnetic bead-based purification of viral nucleic acid.

 
Silica-coated Monarch Mag Beads M1 are utilized for highly sensitive nucleic acid capture from samples containing few target molecules. Silanol groups on the bead shell provide an optimal binding surface for nucleic acids, and the uniform, submicron bead size offers a high surface area with abundant binding sites. Beads are supplied as a monodispersed suspension. The superparamagnetic properties of the bead core result in a fast magnetic response, contributing to ease of handling during use and compatibility with automation

An optimized viral nucleic acid extraction procedure employs a sample lysis step followed by a simple bind-wash-elute process. Samples are treated with Proteinase K and then mixed with a lysis buffer/bead mixture containing lysis buffer, carrier RNA, isopropanol, and magnetic beads, which promote binding of viral nucleic acid onto the silica-coated beads. Carrier RNA is utilized as a co-precipitant in the workflow to enhance the recovery of low amounts of viral nucleic acid. After binding of viral DNA and RNA to the magnetic beads, the beads are washed to remove contaminants, and nucleic acid is eluted in nuclease-free water. Purified viral nucleic acid is suitable for downstream applications, including qPCR/RT-qPCR, ddPCR, and library prep for Next Generation Sequencing (NGS).

Figure 2: Complete SARS-CoV-2 genome coverage from wastewater samples using the Monarch Mag Viral DNA/RNA Extraction Kit

Monarch® Mag Viral DNA/RNA Extraction Kit |
Integrative Genome Viewer visualization of read coverage across the SARS-CoV-2 genome (log scale). RNA was extracted from 750,000 copies of inactivated SARS-CoV-2 viral particles (ATCC® 1986HK) spiked into either the RNA extraction lysis buffer or wastewater samples, using the Monarch Mag Viral DNA/RNA Extraction Kit. Viral particle enrichment was applied to the contrived wastewater samples with Nanotrap® Microbiome A Particles prior to RNA extraction. 20,000 copies of Twist Bioscience Synthetic SARS-CoV-2 RNA Control 23 template served as a positive control for amplicon generation and library prep. Amplicons and libraries were prepared with NEBNext® VarSkip Short v2 SARS-CoV-2 primer pools and the NEBNext ARTIC SARS-CoV-2 FS Library Prep Kit (Illumina). Libraries were sequenced on a NextSeq® 500/550 instrument (2×75 bp). Coverage depth per base was determined, reads were down-sampled with seqtk and aligned to SARS-CoV-2 reference genome (NCBI, NC_045512) with Bowtie2.

Figure 3: Performance comparison of Monarch Mag Viral DNA/RNA Extraction Kit with other suppliers demonstrates high reproducibility and sensitivity of the Monarch kit

Monarch® Mag Viral DNA/RNA Extraction Kit |
Mock samples representing decreasing viral loads were prepared using Heat-inactivated SARS-CoV-2 (ATCC) in VTM (Hardy Diagnostics®). Extraction was performed using Monarch Mag Viral DNA/RNA Extraction Kit and similar kits from two other suppliers. RT-qPCR was performed using NEB #E3019 and BioRad CFX96 Touch Real-Time PCR Detection System. Monarch Mag Viral DNA/RNA Extraction Kit showed consistently low Cts and reproducible data, even at low viral loads, compared to the competitor kits tested.

Figure 4: qRT-PCR on DNA extracted using the Monarch Mag Viral DNA/RNA Extraction Kit demonstrates accurate detection and quantification of target DNA viruses
Monarch® Mag Viral DNA/RNA Extraction Kit |
Monarch Mag Viral DNA/RNA Extraction Kit was used to extract DNA from i. Human Adenovirus- Type 3 (ZeptoMetrix) and ii. Cytomegalovirus (ZeptoMetrix®) at two different viral loads using the automated KingFisher protocol. The eluted DNA was subjected to qRT-PCR on a Bio-Rad® CFX96 Touch™ Real-Time PCR Detection System. Amplification curves demonstrate successful extraction of viral DNA with expected signal corresponding to the respective viral loads.

Figure 5: RT-qPCR on RNA extracted using the Monarch® Mag Viral DNA/RNA Extraction Kit demonstrates detection of multiple viruses from the same sample

Monarch® Mag Viral DNA/RNA Extraction Kit |

Mock co-infection samples representing high and low viral loads were prepared using a swab containing inactivated Influenza and SARS-CoV-2 viruses (Microbiologics®) resuspended in VTM (Hardy Diagnostics®). Extraction was performed using Monarch® Mag Viral DNA/RNA Extraction Kit following the automated KingFisher® protocol. RT-qPCR was performed using relevant primer probes and Bio-Rad® CFX96 Touch™ Real-Time PCR Detection System. Monarch Mag Viral DNA/RNA Extraction Kit can successfully extract nucleic acids from multiple viruses present in a sample.

产品类别:
Nucleic Acid Purification Products

应用:
Nucleic Acid Purification

  • 试剂盒组成

    本产品提供以下试剂或组分:

    NEB # 名称 组分货号 储存温度 数量 浓度
    • T4010S     25    
        Monarch® Mag Beads M1 T4005-21 25 1 x 2.4 ml Not Applicable
        Monarch® Carrier RNA T4006-21 25 1 x 125 µg Not Applicable
        Monarch StabiLyse™ DNA/RNA Buffer T2111-1 25 1 x 24 ml Not Applicable
        Monarch® Buffer BX T2041-1 25 1 x 19 ml Not Applicable
        Nuclease-free Water B1500-2 25 1 x 25 ml Not Applicable
        Proteinase K, Molecular Biology Grade P8200AAVIAL -20 1 x 0.55 ml 20 mg/ml
    • T4010L     25    
        Monarch® Mag Beads M1 T4005-1 25 1 x 14 ml Not Applicable
        Monarch® Carrier RNA T4006-1 25 1 x 750 µg Not Applicable
        Monarch StabiLyse™ DNA/RNA Buffer T2111-3 25 1 x 145 ml Not Applicable
        Monarch® Buffer BX T2041-3 25 1 x 122 ml Not Applicable
        Nuclease-free Water B1500-3 25 1 x 122 ml Not Applicable
        Proteinase K, Molecular Biology Grade P8200SVIAL -20 3 x 1 ml 20 mg/ml
        Proteinase K, Molecular Biology Grade P8200AAVIAL -20 1 x 0.55 ml 20 mg/ml
    • T4010X     25    
        Monarch® Mag Viral DNA/RNA Extraction Kit T4010L 25    
        Monarch® Mag Beads M1 T4005-1 25 3 x 14 ml Not Applicable
        Monarch® Carrier RNA T4006-1 25 3 x 750 µg Not Applicable
        Monarch StabiLyse™ DNA/RNA Buffer T2111-3 25 3 x 145 ml Not Applicable
        Monarch® Buffer BX T2041-3 25 3 x 122 ml Not Applicable
        Nuclease-free Water B1500-3 25 3 x 122 ml Not Applicable
        Proteinase K, Molecular Biology Grade P8200SVIAL -20 9 x 1 ml 20 mg/ml
        Proteinase K, Molecular Biology Grade P8200AAVIAL -20 3 x 0.55 ml 20 mg/ml

  • 相关产品

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操作说明、说明书 & 用法

  • 操作说明

    1. Reagent Preparation Guidance
    2. Protocol for Manual Isolation of Viral DNA/RNA in Microfuge Tubes (1.5 or 2.0 ml)
    3. Protocol Guidance for Automated Isolation of Viral DNA/RNA in Deep Well Plates
    4. Protocol for KingFisher Flex Automated Isolation of Viral DNA/RNA

  • 说明书

    产品说明书包含产品使用的详细信息、产品配方和质控分析。

    • manualT4010

FAQs & 问题解决指南

  • FAQs

    1. The enzymes in the kit were not stored at -20°C right away. Will they still work?
    2. How do I prepare Viral DNA/RNA Wash Buffer?
    3. Does the kit include Nuclease-Free Water to prepare all wash buffers?
    4. Where can I find automation scripts?
    5. What is the sample input volume range?
    6. What sample types are compatible with T4010?
    7. What downstream applications are compatible with T4010?
    8. What is the purpose of Carrier RNA, and is it necessary to include in the extraction?

MABI-Mag200 Magnetic stand for 8-strip PCR tube


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