Ligation, and the 3 accompanying purification steps in the same tube

he Omni-Tube PCR system is designed and optimized for performing PCR, digestion, ligation, and purification all in a single tube. No extra tubes, spin-columns, filter plates, silica membrane, magnetic beads, vacuum or filtration steps are needed for purification, and no transfer of reaction mixtures to different tubes required when proceeding to other downstream applications. Based on Solid Surface Reversible Binding (SSRB) technology, the Omni Tube utilizes plastics coated with proprietary high efficiency-binders acting to selectively capture and efficiently bind DNA fragments (e.g. PCR products, digestions, ligations etc...) from reaction mixtures. In the presence of Binding Buffer, PCR amplicons, or other types of DNA, will specifically interact with the high efficiency-binders and bind to the wells while primers, fluorescent dyes, nucleotides, and other contaminants will remain in solution. Box 1 | Contents
Kit Contents
# of Purifications # of Omni-Tubes Binding Buffer Washing Buffer Elution Buffer ABP-PP-OMNIPCR25 ABP-PP-OMNIPCR50

Omni-Tube PCR, PCR Reaction,Digest,

fter washing the tube to remove unbound material, the purified DNA products can easily be eluted in 10 mM Tris Elution Buffer or water. The purified DNA products can be processed directly in the binding tube for downstream applications such as restriction enzyme digestion, cloning, SNP analysis, and sequencing without transfer to another tube. Everything from PCR down to transformation, with purification steps in between, performed in one Omni Tube!

Features
Fast to perform: Quick 15 minute purification protocol. Easy to handle: Several successive steps in a workflow can be performed in the same tube, e.g. PCR, PCR purification, and a downstream application such as ligation, endonuclease digestion, or sequencing, to mention just a few, without a need for sample transfer. Since no silica membrane is needed for binding, multiple steps of filtration in the sample binding and washing steps associated with conventional silica membrane methods are eliminated, which makes the whole procedure very easy to perform. Reliable quality: Consistent performance, reliable cross-contamination prevention procedure and unique solid-surface capture technology ensure high reproducibility and maximum recovery of high-quality DNA without contamination.

25 25 1.5 mL 3.5 mL 2.5 mL

50 50 3.0 mL 7.0 mL 5.0 mL

Additional Materials Needed 96-100 % ethanol Isopropanol Benchtop Centrifuge

Box 2 | All in 1 Omni-Tube Overview

Proceed to transformation or other downstream applications. Purify ligation

Set up PCR Cocktail in Omni-Tube

Surface Bind technology allows for purification of PCR product in same Omni-Tube.

Purify digest in same Omni-Tube using same protocol.

Ligation (if necessary)

Run PCR using same Omni-Tube

Proceed to digestion reaction (if necessary)

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General Precautions
This kit is for research use only. All due care and attention should be exercised in the handling of the kits. Wear a laboratory coat, disposable gloves, and eye protection when handling reagents and tubes. Avoid ingestion and inhalation of reagents. In case of contact, wash thoroughly with water. See Material Safety Data Sheets (MSDS) for emergency procedures in case of accidental contact or ingestion. MSDS information is available upon request. Always use proper aseptic techniques to avoid nuclease contamination when working with DNA. Use only sterile, new pipette tips to prevent cross contamination. 4. Remove the liquid from the SurfaceBind tube. Choose one of the methods listed below to remove the liquid: (A) Decant the solution by quickly flipping the tube over a waste container and shaking briskly, then place the inverted plate on a stack of clean absorbent paper, such as Kimwipe or paper towels, and tap the tube on the clean paper 3 4 times to remove as much liquid as possible; (B) Remove the solution by aspirating the solution from each well with a multi-channel pipettor. Be sure not to scrape the walls of the wells during aspiration as the products are bound to the walls. Washing DNA Products 1. Add 150 l Wash Buffer containing ethanol to each well. Mix by pipetting solution up and down 2 3 times and incubate the tube for 30 seconds at room temperature. 2. Remove the Wash Buffer from the Omni-Tube using one of the methods suggested in step 4 above. 3. (Optional) Repeat steps 1 and 2 above for a total of two washes. *One wash is sufficient for most applications 4. After the final wash blot the inverted tube dry on a piece of clean absorbent paper 3 4 times and air-dry the tube for 5 8 minutes to remove any residual liquid. Eluting DNA Products
PCR amplicons attached to the wall of the Binding Plate
can be analyzed directly in the well they were purified in by restriction enzyme digestion, primer extension, SNP detection and sequencing using a 25 50 l reaction volume without elution. If the DNA is to be eluted, use the procedure below

Preparation
WB2: Add 100 % ethanol to Washing Buffer II (WB2) and mix well. (Mark bottle that ethanol has been added). Store at room temperature and use WB2 containing ethanol within six (6) months. KB5: Prepare fresh working Binding Buffer (KB5) prior to performing PCR product purification based on the number of samples processed. To make working Binding Buffer (KB5), mix 10 l of Binding Buffer (KB5) with 40 l 100 % Isopropanol and mix well. For working Binding Buffer (KB5), dispense 20 l of Binding Buffer containing Isopropanol per 10-l PCR solution. Discard the unused Binding Buffer at the end of the day. The kits are designed to purify PCR fragment sizes of greater than 60 base pairs from PCR reactions. After PCR,cool the reaction to room temperature before starting the purification.

Protocols
The protocol below is for the purification of PCR DNA amplicons from 50-l PCR using the Omni-Tube PCR system. The purification procedure may be scaled from 10 100 l by proportionately adjusting all reagents throughout the procedure

Binding DNA Products 1. PCR can be performed directly in the OmniTube. If PCR was performed in a different tube, transfer 50 l of PCR mixture from the PCR tube to the Omni-Tube 2. Add 100 l of working Binding Buffer (KB5) containing isopropanol to each sample well. Mix well by pipetting solution up and down 7 8 times. 3. Centrifuge at >2,250 x g for 1 min.

1. Add 50 L Elution Buffer (EB1) into the OmniTube. (If higher concentration is preferred, add 25 L Elution Buffer (EB1) and use Option 1 to elute PCR product). 2. Choose one of the following Options: (Option 1): (Elute with vortex): Vortex the tube for 30 seconds. Centrifuge the tube at maximum speed for 1 minute to collect all liquid at the bottom of each well. (Option 2): (Elute without vortex): Pipette the solution up and down 5 7 times, seal the tube and incubate for 60 seconds at room temperature. 3. The eluted PCR sample can be used immediately for downstream application. Alternatively, the eluted PCR sample may be stored in the sealed plate at 4 C for short-term storage or 20 C for long-term storage.

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Box 2 | Troubleshooting Problem No PCR product Cause PCR failure Solution Be sure to add all components of the PCR mixture. Check PCR positive control.

Low PCR yield Low yield of product Short centrifugation

Check yield of unpurified PCR product on agarose gel.

If lower speed centrifuge is used, extend the spin time.

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