Team:HKUST/Protocols/Agarose gel

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'''Agarose gel preparation and gel electrophoresis''' 

*Purpose: To check the result
*Materials: Agarose, 0.5X TBE, SYBR safe DNA gel stain, DNA loading dye, DNA ladder/marker 
**Choice of ladders: VC100bp Plus DNA ladder (highest band 3000bp); Lamda DNA BstEII marker (highest band 8400bp, 1% argarose)
*Procedure: 
**To make a 0.8% agarose gel, use 0.16 g agarose per 20 mL 0.5x TBE. 
**Cover the flask with plastic wrap to prevent boiling over, then microwave the solution for 1-2 minutes to   dissolve the agarose. 
**Let agarose solution cool for 5 minutes, then add SYBR safe DNA gel stain to a final concentration of 0.5 μg/mL. 
**Pour the agarose solution into a casting tray with well comb in place. Allow 20-30 minutes to completely solidify.
**Place the agarose gel into the gel box and fill the box with 0.5X TBE until the gel is immersed.
**Load an appropriate molecular weight ladder into the first lane of the gel. If needed, an additional ladder with different molecular weight could be loaded into the last lane.
**Add 1 μL loading dye per 5 μL of sample.
**Load the samples from left to right.
**Cover the gel box and plug in the electrodes in a way that the samples will run towards the positive, red electrode.
**Run the gel at 100V until the dye line is approximately 50-75% of the way down the gel. Approximately 20-30 minutes.
**Carefully remove the gel from the gel box and check the result under UV exposure.
*Tips: 
**Higher concentration of agarose solution makes better resolution for less molecular weight expected band.
**Let bottom of the flask be immersed in a cup of cold water for faster cooling.
**In order for clearer band to cut specific band on the gel, immerse the gel in Ethidium bromide for 10-15 minutes after step k.
*Safety tips: 
**Be sure to wear a glove before treating the hot flask.
**Ethidium bromide is a strong mutagen. Wear a lab coat, eye protection, and gloves when working with this chemical.



<ul>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Agarose_gel" >Agarose gel preparation and gel 
electrophoresis</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/PCR">PCR (Taq, Vent)</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/PCR_cleanup">PCR product clean-up</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Enzyme_digestion"> Enzyme digestion (vector, insert)</a></li>			
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/plasmid_extraction"> Plasmid DNA extraction</a></li>	
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/gel_cut"> Gel cutting</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/gel_extraction"> Gel extraction</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Ethanol_precipitation"> Ethanol precipitation</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Ligation"> Ligation</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Transformation_to_Ecoli"> Transformation to E.coli</a></li>
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yeast</a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Yeast_genomic_DNA_extraction"> Yeast genomic DNA 
extraction</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Western_blotting"> Western blotting</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/GFP_testing"> GFP testing</a></li>
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Biological_test"> Biological test</a></li>
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