Protocol

One-Step PCR Inhibitor Removal Kit

Fast & Effective Removal of PCR Inhibitors

Products

BcMag™ One-Step PCR Inhibitor Removal Kit

Cat. No. AX101

Unit Size 1 ml

Order

BcMag™ One-Step PCR Inhibitor Removal Kit

Cat. No. AX102

Unit Size 5 ml

Order

Previous slide

Next slide

Please contact us for a quote on larger sizes …

Cat. No.

AX101

AX102

Product Name

BcMag™ One-Step PCR Inhibitor Removal Kit

Unit Size

1 ml

5 ml

Order

Please contact us for a quote on larger sizes …

Store at 4°C upon arrival for up to 6 months.

Description

Purification Kit

Instruction Manual

MSDS

Introduction

PCR technology has become one of the most potent molecular biological tools in the last few decades. However, biological samples collected from different materials often contain various PCR inhibitors that can interfere with PCR amplification. PCR inhibitors are a diverse group of compounds that inhibit the amplification of DNA through the polymerase chain reaction (PCR). PCR inhibitors generally exert their effects by directly binding the active site of a DNA polymerase to cause decreased sensitivity or complete failures of the DNA amplification. Therefore, removing PCR inhibitors from the DNA extracts before the PCR amplification is vital for all downstream applications.

BcMag™ One-Step PCR Inhibitor Removal Kit provides one-step removal of PCR inhibitor from impure DNA samples before PCR, RT, and other downstream applications based on negative chromatography. The magnetic beads are superparamagnetic and modified with our proprietary chemistry. When mixed with inhibitor-containing samples, the beads instantly capture and remove the PCR inhibitors. At the same time, only the pure DNA remains in the solution and is ready for all downstream applications. The beads can effectively remove many common inhibitors such as polyphenolic compounds, humic/fulvic acids, acidic polysaccharides, tannins, melanin, heparin, detergents, and denim dyes, and divalent cations such as Ca²⁺, Mg²⁺, etc.

Workflow

The protocol is straightforward and fast: one tube, one step, and one minute (Fig.1). Add the magnetic beads directly to the pre-purified DNA samples and vortex or pipette to capture and remove the impurities. After vortexing/pipetting, the beads are magnetically removed, while the supernatant contains the purified and ready-to-run products. Unlike standard bind-wash-elute protocol, this convenient procedure does not contain traces of organic solvents, chaotropic salts, or EDTA and is almost 100% DNA recovery. The beads enable 96 samples to be processed simultaneously in less than 10 minutes with cost-effective lab vortex mixers.

Features and Advantages

●

Simple Protocol: No liquid transfer, One-tube, One-step

●

Ultrafast: One-minute manual protocol or less than 10-minutes vortex (96 samples)

●

Higher purity and recovery > 90% DNA (> 50bp)

●

Effective Cleanup: polyphenolic compounds, humic/fulvic acids, acidic polysaccharides, tannins, melanin, detergents, divalent cations such as Ca²⁺, Mg²⁺, etc.

●

Cost-effective: Eliminates columns, filters, laborious repeat pipetting, and ethanol

●

High throughput: Compatible with many different automated liquid handling systems

Handling and Storage:

Store at 4°C upon arrival for up to 12 months.

DO NOT FREEZE

PROTOCOL

The following protocol is an example. The beads and sample volume can be rational Scale-up (or down) as needed. Do not use buffers containing organic solvents.

Equipment

Item

Magnetic Rack for centrifuge tube

** Based on sample volume, the user can choose one of the following magnetic Racks

Source

• BcMag™ Rack-2 for holding two individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-01)

• BcMag™ Rack-6 for holding six individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-02)

• BcMag™ Rack-24 for holding twenty-four individual 1.5-2.0 ml centrifuge tubes (Bioclone, Cat. No. MS-03)

• BcMag™ Rack-50 for holding one 50 ml centrifuge tube, one 15 ml centrifuge tube, and four individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-04)

Item

BcMag™ 96-well Plate Magnetic Rack.

Source

• BcMa™ 96-well Plate Magnetic Rack (side-pull) compatible with 96-well PCR plate and 96-well microplate or other compatible Racks (Bioclone, Cat. No. MS-06)

Item

Adjustable Single and Multichannel pipettes

Item

Centrifuge with swinging bucket

Addition items are required if using 96-well PCR plates / tubes

Vortex Mixer

** The user can also use other compatible vortex mixers. However, the Time and speed should be optimized, and the mixer should be: Orbit ≥1.5 mm-4 mm, Speed ≥ 2000 rpm

Eppendorf™ MixMate™

Eppendorf, Cat. No. 5353000529

Tube Holder PCR 96

Eppendorf, Cat. No. 022674005

Tube Holder 1.5/2.0 mL, for 24 × 1.5 mL or 2.0 mL

Eppendorf, Cat. No. 022674048

Smart Mixer, Multi Shaker

BenchTop Lab Systems, Cat. No. 5353000529

1.5/2.0 mL centrifuge tube

96-well PCR Plates or 8-Strip PCR Tubes

PCR plates/tube

! IMPORTANT ! If using other tubes or PCR plates, make sure that the well diameter at the bottom of the conical section of PCR Tubes or PCR plates must be ≥2.5mm.

Addition Items Are Required If Using 96-Well Microplates

Fisher Scientific™ Microplate Advanced Vortex Mixers

Fisher, Cat. No. 02-216-101

OHAUS Microplate Vortex Mixers

OHAUS, Cat. No. 30392160

Vortex Mixer

** The user can also use other compatible vortex mixers. However, the time and speed should be optimized, and the mixer should be Orbit ≥1.5 mm-4 mm, Speed≥ 800 rpm

Clear Flat-Bottom Non-Binding Assay Microplates

Items

Magnetic Rack for centrifuge tube

** Based on sample volume, the user can choose one of the following magnetic Racks

Source

●

BcMag™ Rack-2 for holding two individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-01)

●

BcMag™ Rack-6 for holding six individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-02)

●

BcMag™ Rack-24 for holding twenty-four individual 1.5-2.0 ml centrifuge tubes (Bioclone, Cat. No. MS-03)

●

BcMag™ Rack-50 for holding one 50 ml centrifuge tube, one 15 ml centrifuge tube, and four individual 1.5 ml centrifuge tubes (Bioclone, Cat. No. MS-04)

BcMag™ 96-well Plate Magnetic Rack

●

BcMa™ 96-well Plate Magnetic Rack (side-pull) compatible with 96-well PCR plate and 96-well microplate or other compatible Racks (Bioclone, Cat. No. MS-06)

Adjustable Single and Multichannel pipettes

Centrifuge with swinging bucket

Addition items are required if using 96-well PCR plates/tubes

Vortex Mixer

** The user can also use other compatible vortex mixers. However, the Time and Speed should be optimized, and the mixer should be: Orbit ≥1.5 mm-4 mm, Speed ≥ 2000 rpm

Eppendorf™ MixMate™

Tube Holder PCR 96

Tube Holder 1.5/2.0 mL, for 24 × 1.5 mL or 2.0 mL

Smart Mixer, Multi Shaker

Eppendorf, Cat. No. 5353000529

Eppendorf, Cat. No. 022674005

Eppendorf, Cat. No. 022674048

BenchTop Lab Systems, Cat. No. 5353000529

Eppendorf™ MixMate™

Tube Holder PCR 96

Tube Holder 1.5/2.0 mL, for 24 × 1.5 mL or 2.0 mL

Smart Mixer, Multi Shaker

Eppendorf, Cat. No. 5353000529

Eppendorf, Cat. No. 022674005

Eppendorf, Cat. No. 022674048

BenchTop Lab Systems, Cat. No. 5353000529

1.5/2.0 mL centrifuge tube

96-well PCR Plates or 8-Strip PCR Tubes

PCR plates/tubes

! IMPORTANT ! If using other tubes or PCR plates, make sure that the well diameter at the bottom of the conical section of PCR Tubes or PCR plates must be ≥2.5mm.

Addition items are required if using 96-well microplates

Fisher Scientific™ Microplate Advanced Vortex Mixers

OHAUS Microplate Vortex Mixers

Fisher, Cat. No. 02-216-101

OHAUS, Cat. No. 30392160

Fisher Scientific™ Microplate Advanced Vortex Mixers

Fisher, Cat. No. 02-216-101

OHAUS Microplate Vortex Mixers

OHAUS, Cat. No. 30392160

Vortex Mixer

** The user can also use other compatible vortex mixers. However, the time and speed should be optimized, and the mixer should be Orbit ≥1.5 mm-4 mm, Speed≥ 800 rpm

Clear Flat-bottom Non-Binding Assay Microplates

Procedure

Transfer 100µl premix beads solution (Table 2) to the sample to a new well of 96well PCR plate or 0.2ml PCR tube and add the sample.

! IMPORTANT ! It is essential to mix the beads before dispensing. Do not allow the beads to sit for more than 2 minutes before dispensing. Resuspend the magnetic beads every 2 minutes.

Add 10µl magnetic beads to a 100 µl pre-purified DNA sample.

! IMPORTANT ! Users need to optimize the ratio of beads, and the concentration of the PCR inhibitors since the concentration of the inhibitors varies from sample to sample.

Mix the sample with beads for 1 minute by slowly pipetting up and down 20-25 times or by vortex mixer at 2000 rpm for 5 minutes.

Place the sample plate or tube on the magnetic separation plate for 30 seconds or until the solution is clear.

Transfer the supernatant to a clean plate /tube while the sample plate remains on the magnetic separation plate. The sample is ready for downstream applications.

Learn More

Instruction Manual

MSDS

DNA & RNA Purification Related Products →

General Reference

Poddar SK, Sawyer MH, Connor JD. Effect of inhibitors in clinical specimens on Taq and Tth DNA polymerase-based PCR amplification of influenza A virus. J Med Microbiol. 1998 Dec;47(12):1131-5.

Kaltenboeck B, Wang C. Advances in real-time PCR: application to clinical laboratory diagnostics. Adv Clin Chem. 2005;40:219-59.

Alaeddini R. Forensic implications of PCR inhibition – A review. Forensic Sci Int Genet. 2012 May;6(3):297-305.

Al-Soud WA, Rådström P. Purification and characterization of PCR-inhibitory components in blood cells. J Clin Microbiol. 2001 Feb;39(2):485-93.

Bioclone

Protocol

One-Step PCR Inhibitor Removal Kit

Fast & Effective Removal of PCR Inhibitors

Introduction

Workflow

Features and Advantages

PROTOCOL

Learn More

General Reference

Bioclone Inc.

Welcome to BitClone