From 2011.igem.org
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| - | {{:Team:DTU-Denmark/Templates/Standard_page_begin|Plasmid purification}}
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| - | == Plasmid purification by miniprep (Zymo Research Group) ==
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| - | # Spin down 2 ml of cell culture in a 2 ml Eppendorf tube at 11.000g for 5 min
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| - | # Remove supernatant, spin again down for 10 seconds, remove supernatant (if the pellet is not sufficient, repeat step 1 in the same tube)
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| - | # Resuspend your pellet in 600 ul TE buffer
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| - | # Add 100 µl 7X Lysis Buffer (blue) and mix by inverting the tube 4-6 times. Proceed to the next step within 2 minutes.
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| - | # Add 350 µl cold Neutralization Buffer (Yellow) and mix carefully. The sample will turn yellow and a yellowish precipitate will occur, then the reaction is finished. Invert the tube an additional 2-3 times to ensure complete neutralization.
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| - | # Centrifuge at 11,000 g for 5 min.
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| - | # Transfer the supernatant (ca. 875 µl) into a column. Avoid disturbing the cell debris pellet.
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| - | # Place the column into a collection tube and centrifuge for 30 seconds.
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| - | # Discard the flow-through and place the column back into the same collection tube.
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| - | # Add 200 µl Endo-Wash-Buffer to the column and centrifuge for 30 seconds.
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| - | # Add 400 µl of ZyppyWash Buffer to the column and centrifuge for 30 seconds.
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| - | # Transfer the column into a clean 1,5 ml Eppendorf tube and then add 50 µl Zyppy Elution Buffer directly to the column. Let the product(s) stand for 1-15 minutes on the table at RT.
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| - | # Centrifuge for 30 seconds to elute the plasmid DNA.
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| - | {{:Team:DTU-Denmark/Templates/Standard_page_end}}
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Latest revision as of 19:17, 21 September 2011
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