ISOSPIN PCR Product
Manual (Ver. 03)

Code No. 315-08001

NIPPON GENE CO., LTD.

IDescription

The ISOSPIN PCR Product enables a rapid and effective purification of DNA fragments fromamplification reactions.

II Kit components
Component(100 preps)Note

ISB Buffer

100 ml x 1

ISW Buffer

100 ml x 1(includes ethanol)*

ISE Buffer

10 ml x 1

10 mM Tris-HCl (pH 8.5)

Spin Column (a Spin Column and Collection Tube set) 50 sets x 2*III

Keep the buffer bottle tightly closed after use.

Storage conditions

All the kit components can be stored at room temperature (15-25°C).

1IV Precautions
・ The kit is intended research use only.

V Protocol
Material not supplied
・Micropipette・Pipette tips
・1.5 mL microcentrifuge tubes
・Microcentrifuge

2Protocol1.

Add 1 volume of Sample (PCR reaction) in a 1.5 mL microcentrifuge tube.
Add 5 volumes of ISB Buffer, and mix well by inverting. Spin down lightly.
Example) Add 250 µl of ISB Buffer to 50 µl of the Sample.

2.

Prepare a Spin Column (a Spin Column and Collection Tube set). Apply theMixture to the Spin Column.

3.

Centrifuge for 1 min at 12,000 x g at room temperature.

4.

Discard the flow-through. Place the Spin Column back into the same
Collection Tube.
Note) If there are the Mixture left over, apply the remaining Mixture to the Spin Column, andrepeat 3 and 4.

5.

Apply 750 µl of ISW Buffer to the Spin Column.

6.

Centrifuge for 1 min at 12,000 x g at room temperature.

7.

Discard the flow-through. Place the Spin Column back into the same
Collection Tube.

8.

Centrifuge for 1 min at 12,000 x g at room temperature to remove residualliquid.

9.

Transfer the Spin Column to a new 1.5 mL microcentrifuge tube.

10.

Add 50 µl of ISE Buffer to the center of the Spin Column. Incubate 3 min atroom temperature.
Note) Nuclease-free water or TE (pH8.0) can also be used to elute the DNA in place of ISE
Buffer (10 mM Tris-HCl, pH 8.5).

11.

Centrifuge for 1 min at 12,000 x g at room temperature.

12.

Recover your purified DNA fragments in the microcentrifuge tube.
Note) The DNA fragments can be used directly or stored at -20°C for long term storage.

3Simplified Protocol
Sample in a 1.5 mL microcentrifuge tube
Add 5 volumes of ISB Buffer to 1 volume of Sample.
Mix by inverting.
Spin down lightly.
Apply the Mixture to a Spin Column.
Centrifuge for 1 min at 12,000 x g at room temperature.
Discard the flow-through.
Add 750 µl of ISW Buffer.
Centrifuge for 1 min at 12,000 x g at room temperature.
Discard the flow-through.
Centrifuge for 1 min at 12,000 x g at room temperature to remove residual liquid.
Transfer the Spin Column to a new 1.5 mL microcentrifuge tube.
Apply 50 µl of ISE Buffer to the center of the Spin Column.
Incubate 3 min at room temperature.
Centrifuge for 1 min at 12,000 x g at room temperature.
Recover your purified DNA fragments in the microcentrifuge tube.

4VI

Troubleshooting

Low DNA Yield
Ensure the ISE Buffer is applied to the center of the Spin Column so that elution is efficient.
Larger elution volumes can increase yield of DNA at the cost of dilution of the sample.

Low DNA Performance
Please repeat wash step (Step 5, 6 and 7). 80% Ethanol can also be used to wash the DNA inplace of ISW Buffer.
If ethanol has been carried-over, spin for 5 min, instead of 1 min in Step 8.

Sheared DNA
Mix well by inverting or flicking the tube. Do not vortex.

5VII

Data

Technical Information
Binding Capacity:

20 µg

Minimum Elution Volume:

10 µl

Typical DNA Recovery:

60–95% (100 bp to 20 kbp)

Column Volume:

900 µlRemovable Primer Size:≤ 40 mer

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The information in the descriptions of the products may be changed without prior notification.

NIPPON GENE CO., LTD.
If you have any questions, please contact us by web form.

http://www.nippongene.com/ISOSPIN PCR Product Manual (Ver. 04) 201311-1704

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