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Team:Sumbawagen/Notebook/protocol4
From 2014.igem.org
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<p><u>DNA Purification from Liquid Samples</u></p> | <p><u>DNA Purification from Liquid Samples</u></p> | ||
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| - | <p>1. Into a clean, | + | <p>1. Into a clean, sterillized 0.5 ul polypropylene tube, add PCR products : buffer GP1 = 1 : 5 (e.g. 40 ul : 200 ul). Vortexing.</p> |
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p> | <p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p> | ||
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p> | <p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p> | ||
Revision as of 01:50, 18 October 2014
Team:Sumbawagen/Team
From 2014.igem.org
Notebook – Protocol – 4. DNA Purification
Spin column based purification was used. The kit is Gel/PCR extraction kit (Nippon Genetics). Protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge. All steps were done at room temperature.
DNA Purification from Liquid Samples
1. Into a clean, sterillized 0.5 ul polypropylene tube, add PCR products : buffer GP1 = 1 : 5 (e.g. 40 ul : 200 ul). Vortexing.
2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.
3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.
4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.
5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube.
6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.
7. Obtain the DNA as the flowthrough.
DNA Purification from Agarose Gel
1. Into a clean, sterlized 1.5 ul polypropylene tube, add cutted gel containing DNA of interest : buffer GP1 = < 300 mg : 500 ul. Vortexing. Incubate at 55 oC; 10-15 minutes. Invert the tube.
2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.
3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.
4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.
5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube
6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.
7. Obtain the DNA as the flowthrough.
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