LaboPassTMPCR

Protocol Book

LaboPass TM PCR
Introduction
LaboPass TM PCR kit provides a simple and rapid method to purify PCR
products or other enzymatic reactions in just 6 minutes. Up to 10 µg of pure
DNA which is at least 100 bp but less than 50 kb in length can be obtainedusing this kit and the purified DNA can be directly used for sequencing,cloning and other routine applications without further manipulation.
LaboPass TM PCR procedures remove the DNA fragments smaller than 100
bp from the preparation, resulting in removal of primers or primer dimers ofPCR.

Principle of methods
In the presence of high salt, DNA binds selectively to a glassfiber membrane
in a LaboPass TM spin column. The binding reaction occurs due to thedisruption of the organized structure of water molecules and the interaction
with the nucleic acid. The DNA remains bound while a series of rapid "washand-spin" steps removes contaminating small molecules (primers, nucleotideand salts). Finally, a low salt buffer from the glassfiber membrane elutes thepurified DNA. The process does not require DNA precipitation, organicsolvent extractions, or further handling.

Storage and Stability
LaboPass TM PCR kit is shipped at room temperature. All solutions should be
stored at 15 – 25℃. The kit components are guaranteed to be stable for 12months from the date of manufacture printed on the product package.
Please note that improper storage at 4℃ (refrigerator) or -20℃ (freezer) will
adversely impact DNA purification when precipitates form in the buffers. The
Buffer PB may exhibit salt precipitation due to cold temperatures as shippingor lab ambient conditions in winter. The procedure will not be performed
optimally if the salt precipitates out of solution. If that happen in any of thesolutions, warm the bottle at 50℃ with occasional mixing until completelydissolved.

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Chemical Hazard
Buffer PB contains irritant which is harmful when in contact with skin or eyes,
or when inhaled or swallowed. Care should be taken during handling.
Always wear gloves and eye protector, and follow standard safety
precautions.

Quality Control
All components in this kit are manufactured in particle-controlled area(class1000 clean room), and its degree of cleanness is monitored periodically.

Kit Contents
Cat. No.
No. of preparation
Spin column
Collection tube
Buffer PB
Buffer NW(conc.)
Buffer EB

CMR0111
5050
50
30ml
15ml
15ml

CMR0112
200
200
200
125ml
57ml
15ml

CMR0115
1000
1000
1000
125mlx557mlx515mlx5* All Solution (Buffer) is available to purchase separately.

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LaboPass TM PCR kit Protocols
Before experiment- All centrifugation should be carried out at 10,000 xg above (>12,000 rpm) at room

temperature in a microcentrifuge.

- Add absolute ethanol to Buffer NW as printed on the label before
first use.
- All solutions should be equilibrated at room temperature before procedures.

1. Add 5 volumes of Buffer PB to 1 volume of the sample and mix.
Transfer the mixture to a Spin Column.
For 100 µl reaction, add 500 µl of Buffer PB. It is not necessary to remove
mineral oil. If precipitated material has formed in buffer PB, heat to dissolve
at 37℃.

2. Centrifuge for 1 min. Discard the flow through and reinsert theSpin Column back into the same tube.
3. Add 750µl of Buffer NW. Centrifuge for 1 min at full speed.
Transfer the Spin Column into a new 1.5 ml tube.
If the Spin Column has Buffer NW associated with it, centrifuge again for
additional 1 min at full speed before transferring to a new 1.5 ml tube.
Residual ethanol from buffer NW may inhibit subsequent enzymatic reaction.

4. Apply 50µl of Buffer EB or dH2O to the center of the membranein the Spin Column, let stand for 1 min, and centrifuge for 1 min.
Elution volume can be reduced to 30 µl for higher concentration of elute.
Ensure that the buffer EB or distilled water is dispensed directly onto thecenter of spin column membrane for optimal elution of DNA. For long-term
storage, eluting in buffer EB (10mM Tris-Cl. pH 8.5) or TE, pH 8.0 andstoring at –20℃ is recommended. When using water for elution, check thepH of water (>pH7.0) before elution.

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Troubleshooting guide
Low or no recovery
Buffer NW prepared incorrectly- Ensure that the appropriate volume of absolute ethanol printed on the bottle label
is added to Buffer NW.
Improper elution buffer- As user's requirement, elution buffer other than Buffer EB can be used. However,the conditions of optimal elution should be low salt concentration with alkaline pH(7<pH<9). When water or other buffer was used as eluent, ensure that conditions.
Elution buffer incorrectly dispensed- Ensure that elution buffer dispensed to the center of membrane. Incorrectly
dispensed elution buffer causes inappropriate contact with membrane, followed bypoor DNA recovery.
Enzymatic reactions is not performed well with purified DNA
Residual ethanol from Buffer NW remains in eluate- Discard flow-through from step 3, and then centrifuge for additional 2 min toremove ethanol completely before proceed to step 4.
Eluate contains denatured ddDNA- For reannealing of ssDNA to dsDNA, incubate ssDNA at 95℃ for 2 min, and thenallow to cool to room temperature.
DNA floats out of well while loading of agarose gel
Residual ethanol from Buffer NW remains in eluate- Discard flow-through from step 3, and then centrifuge for additional 2 min toremove ethanol completely before proceed to step 4.

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Contact us ;製造元COSMO GENETECH CO.,Ltd.

Internet : www.cosmo4.com
E-mail : LaboPass@cosmo4.com
Tel : 82-2-922-7307
Fax : 82-2-922-7307

販売元
北海道システム・サイエンス株式会社

HokkaidoSystem Science.,Ltd.
北海道システム・サイエンス株式会社〒001-0932
札幌市北区新川西2条1丁目2-1
TEL : 011-768-5901
FAX : 011-768-5951
E-mail : bio@hssnet.co.jp
URL : http://www.hssnet.co.jp/

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