Team:HKUST/Protocols/Yeast genomic DNA extraction

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 Home Our Team Project Description

<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

Yeast genomic DNA extraction

Purpose: To extract yeast genomic DNA from yeast strain (strain YPH501, in our case). Materials: yeast strain, lysis buffer, phenol, chloroform, 3M NaAc, 100% ethanol, 70% ethanol, ddH2O, glass beads.

Procedure: a.Add 200μL lysis buffer. b.Pick a whole patch of colony and add it in the tube. c.Add 100μL phenol, and mix it well. d.Add 100μL chloroform, and a few glass beads. e.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm). f.Put the supernatant (~160μL) to another tube. g.Add 200μL phenol/chloroform (1:1 in volume) h.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm). i.Put the supernatant (~120μL) to another tube. j.Add 0.1 volume NaAc(pH5.2), approximately 12μL. k.Add 2 volume of 100% ethanol, approximately 240μL. l.Store it at -20 °C for 20mins. m.Centrifuge for 10mins (13,000 rpm) and remove the supernatant. n.Add 2 volume of 70% ethanol. o.Centrifuge for 5mins (13,000 rpm) and remove the supernatant. p.Stand the tube without closing it for a while to let the ethanol evaporate. q.Add 20μL ddH2O to resuspend it. r.Store it at -20 °C.

Safety tips: Phenol is a strong toxin, make sure to wear gloves and be careful with this chemical.

<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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