Team:HKUST/Protocols/gel extraction

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<b> Main Parts</li> </b> Odorant Sensing</a></li> Attractant Production</a></li> Toxin Production</a></li>

<b> Resources</li> </b>

Lab Notebook</a></li> Parts Submitted </a></li> Protocol List</a></li> Other Resources</a></li> </ul> <ul> <li><a href="http://2009.igem.org/Team:Gallery">Gallery</a></li> <li><a href="http://2009.igem.org/Team:Biosafety">Biosafety</a></li> <li><a href="http://2009.igem.org/Team:Acknowledgement">Acknowledgement</a></li> </ul> a

Gel extraction

Purpose: To clean up the gel after band cutting from the gel and leave only the desired DNA fragment. Materials: GEX Buffer, WN Buffer, WS Buffer, Elution Buffer (all provided in FavorPrepTM Gel/PCR DNA Clean-Up Kit)

Procedure: a.Add 0.5ml GEX buffer into the tube with gel fragment in it. b.Incubate at 60°C for 5-10mins until the gel is completely dissolved. Invert the tube every 2 mins during the incubation. Stop incubation after the gel is completely dissolved. Let the gel mixture to cool down to room temperature. c.Place a GP column onto a collection tube. Load no more than 0.7ml gel mixture into each column. Centrifuge for 30s-60s (9000rpm). Discard the flow through. d.Repeat step c for the excessive gel mixture. e.Wash the column once with 0.5ml WN buffer by centrifuging for 30s-60s (9000rpm). Discard the flow through. f.Wash the column once with 0.5ml WS buffer by centrifuging for 30s-60s (9000rpm). g.Centrifuge for another 3 mins (13200rpm) to remove all the ethanol h.Place the column onto a new 1.5ml centrifuge tube. Add 15μL -30μL elution buffer onto the center of the membrane. (Preheated the Elution buffer to 60°C). i.Stand the column for another 3mins and then centrifuge at full speed for 1-2 mins to elute DNA. Tips: The gel extraction has a low recovery rate of about 10%. Stand the column longer in step i may give relatively higher recovery rate. <ul> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Agarose_gel" >Agarose gel preparation and gel electrophoresis</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/PCR">PCR (Taq, Vent)</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/PCR_cleanup">PCR product clean-up</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Enzyme_digestion"> Enzyme digestion (vector, insert)</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/plasmid_extraction"> Plasmid DNA extraction</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/gel_cut"> Gel cutting</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/gel_extraction"> Gel extraction</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Ethanol_precipitation"> Ethanol precipitation</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Ligation"> Ligation</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Transformation_to_Ecoli"> Transformation to E.coli</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Transformation_to_yeast">Transformation to yeast</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Cell_lysis"> Cell lysis</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Yeast_genomic_DNA_extraction"> Yeast genomic DNA extraction</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/Western_blotting"> Western blotting</a></li> <li><a href="http://2009.igem.org/Team:HKUST/Protocols/GFP_testing"> GFP testing</a></li>

</ul> iGEM 2009

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