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| | <!-- https://2012.igem.org/Team:EPF-Lausanne/Protocol --> | | <!-- https://2012.igem.org/Team:EPF-Lausanne/Protocol --> |
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| - | 1. Download the following spreadsheet : [[File:Team-EPF-Lausanne Ligation.xls]]
| + | Ligation is a method of combining several DNA fragments into a single plasmid. This is often the |
| | + | step following a [[Team:EPF-Lausanne/Protocol/PCR|PCR]] (and a [[Team:EPF-Lausanne/Protocol/PCRCleanup|PCR cleanup]]) or a [[Team:EPF-Lausanne/Protocol/GelExtraction|gel extraction]]. You can also do a "dirty" ligation, where you follow a certain number of [[Team:EPF-Lausanne/Protocol/RestrictionSiteDigestion|digestions]] directly by a ligation. |
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| - | 2. Fill in the pink areas with the vector and fragment concentration, their size and the ratio.
| + | # Download the following spreadsheet : [[File:Team-EPF-Lausanne Ligation.xls]] |
| | + | # Fill in the pink areas with the vector and fragment concentration, their size and the ratio. |
| | + | # Add all the suggested ingredients order in a microcentrifuge tube, in the order they appear. |
| | + | # Ligate for 2 hours at 14ºC. |
| | + | # Immediately transform competent bacteria with the ligation product. |
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| - | 3. Add all the suggested ingradients in a microcentrifuge tube, in the order they appear.
| + | Note: This protocol hasn't been optimized for blunt-end ligation (though it might still work). |
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| - | 4. Ligate for 2 hours at 14ºC.
| + | |
| - | | + | |
| - | 5. Inmediatly transform competent bacteria with the ligation product.
| + | |
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| | {{:Team:EPF-Lausanne/Template/ProtocolFooter}} | | {{:Team:EPF-Lausanne/Template/ProtocolFooter}} |
| | <noinclude>{{:Team:EPF-Lausanne/Template/Footer}}</noinclude> | | <noinclude>{{:Team:EPF-Lausanne/Template/Footer}}</noinclude> |
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