Team:USTC CHINA/Notebook/Protocols/Concentrating proteins

From 2013.igem.org

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<h1>Concentrating proteins</h1>
<h1>Concentrating proteins</h1>
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<p>Performed with Sangon Biotech(Shanghai)  
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<p>Performed with Sangon Biotech(Shanghai)</br>
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Protocol:
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Protocol:</br>
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1.  Prepare a 1.5ml clean centrifuge tube and add 200ul sample protein solution to the tube.
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1.Prepare a 1.5ml clean centrifuge tube and add 200ul sample protein solution to the tube.</br>
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2.  Add 50ul precipitaton solution A and reverse the tube vertically for 10 seconds to make it mixed up evenly.  
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2.Add 50ul precipitaton solution A and reverse the tube vertically for 10 seconds to make it mixed up evenly. </br>
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3. Put the tube on ice or the substratum of fridge and incubate it for an hour.
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3.Put the tube on ice or the substratum of fridge and incubate it for an hour. </br>
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4.  Centrifuge the tube for 15 minutes with 15000rpm, temperature 4℃.  
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4.Centrifuge the tube for 15 minutes with 15000rpm, temperature 4℃. </br>
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5.  Remove the supernatant and the liquid on the tube wall and tube bottom, and remain the sediment.  
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5.Remove the supernatant and the liquid on the tube wall and tube bottom, and remain the sediment. </br>
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6. Add 600ul washing liquor and reverse the tube vertically for 10 seconds.  
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6.Add 600ul washing liquor and reverse the tube vertically for 10 seconds. </br>
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7.  Put the tube on the substratum of fridge for 10 minutes and centrifuge the tube for 15 minutes with 12000rpm, temperature 4℃ to get the sediment.
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7.Put the tube on the substratum of fridge for 10 minutes and centrifuge the tube for 15 minutes with 12000rpm, temperature 4℃ to get the sediment.</br>
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8.  Pour away the supernatant and dry out the sediment in the draught cupboard for 30 minutes or unwater the sediment with freeze drier.  
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8.Pour away the supernatant and dry out the sediment in the draught cupboard for 30 minutes or unwater the sediment with freeze drier. </br>
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9.  Add 20ul solution C and reverse the tube vertically for 10 seconds; if the solution turns yellow, add 1-5ul buffer solution to make it blue  
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9.Add 20ul solution C and reverse the tube vertically for 10 seconds; if the solution turns yellow, add 1-5ul buffer solution to make it blue </br>
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10. Boiling the solution for 5 minutes and centrifuge it for 5 minutes with 12000rpm at room temperature.
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10.Boiling the solution for 5 minutes and centrifuge it for 5 minutes with 12000rpm at room temperature.</br>
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12. Sample the supernatant and carry out SDS-PAGE electrophoresis and analyze the molecule weight
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12.Sample the supernatant and carry out SDS-PAGE electrophoresis and analyze the molecule weight.</br>

Revision as of 14:51, 25 September 2013

Concentrating proteins

Performed with Sangon Biotech(Shanghai)
Protocol:
1.Prepare a 1.5ml clean centrifuge tube and add 200ul sample protein solution to the tube.
2.Add 50ul precipitaton solution A and reverse the tube vertically for 10 seconds to make it mixed up evenly.
3.Put the tube on ice or the substratum of fridge and incubate it for an hour.
4.Centrifuge the tube for 15 minutes with 15000rpm, temperature 4℃.
5.Remove the supernatant and the liquid on the tube wall and tube bottom, and remain the sediment.
6.Add 600ul washing liquor and reverse the tube vertically for 10 seconds.
7.Put the tube on the substratum of fridge for 10 minutes and centrifuge the tube for 15 minutes with 12000rpm, temperature 4℃ to get the sediment.
8.Pour away the supernatant and dry out the sediment in the draught cupboard for 30 minutes or unwater the sediment with freeze drier.
9.Add 20ul solution C and reverse the tube vertically for 10 seconds; if the solution turns yellow, add 1-5ul buffer solution to make it blue
10.Boiling the solution for 5 minutes and centrifuge it for 5 minutes with 12000rpm at room temperature.
12.Sample the supernatant and carry out SDS-PAGE electrophoresis and analyze the molecule weight.