Team:HKUST/Protocols/Yeast genomic DNA extraction

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(New page: 13.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 Na...)
 
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13.Yeast genomic DNA extraction
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<html xmlns="http://www.w3.org/1999/xhtml">
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   Purpose: To extract yeast genomic DNA from yeast strain (strain YPH501, in our case).
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<head>
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   Materials: yeast strain, lysis buffer, phenol, chloroform, 3M NaAc, 100% ethanol, 70% ethanol, ddH2O, glass beads.
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<meta http-equiv="Content-Type" content="text/html; charset=iso-8859-1" />
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  Procedure:
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<link href="http://igem2009hkust.fileave.com/wiki/template/12092009/style.css " rel="stylesheet" type="text/css" />
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   a.Add 200μL lysis buffer.
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<title>Salt and Soap template</title>
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   b.Pick a whole patch of colony and add it in the tube.
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</head>
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   c.Add 100μL phenol, and mix it well.
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   d.Add 100μL chloroform, and a few glass beads.
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<bodyxx>
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   e.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm).  
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<div id="containerxx">
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   f.Put the supernatant (~160μL) to another tube.  
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<div id="headerxx">
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   g.Add 200μL phenol/chloroform (1:1 in volume)
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   h.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm).  
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</div>
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   i.Put the supernatant (~120μL) to another tube.  
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<div id="borderxx">
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   j.Add 0.1 volume NaAc(pH5.2), approximately 12μL.
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<div id="mainxx">
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   k.Add 2 volume of 100% ethanol, approximately 240μL.
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<div id="leftxx">
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   l.Store it at -20 °C for 20mins.
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<div id="menuxx">
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   m.Centrifuge for 10mins (13,000 rpm) and remove the supernatant.
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<ul>
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   n.Add 2 volume of 70% ethanol.
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<li><a href="https://2009.igem.org/Team:HKUST">Home</a></li>
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   o.Centrifuge for 5mins (13,000 rpm) and remove the supernatant.  
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<li><a href="https://2009.igem.org/Team:HKUST/Team">Our Team</a></li>
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   p.Stand the tube without closing it for a while to let the ethanol evaporate.
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<li><a href="https://2009.igem.org/Team:HKUST/Project">Project Description</a></li>
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   q.Add 20μL ddH2O to resuspend it.
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   r.Store it at -20 °C.
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<b>
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  Safety Tips:
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<span style="color:green">
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  •Phenol is a strong toxin, make sure to wear gloves and be careful with this chemical.
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<li>Main Parts</li>
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</span>
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</b>
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<li><a href="https://2009.igem.org/Team:HKUST/OdorantSensing">Odorant Sensing</a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/AttractantProduction">Attractant Production</a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/ToxinProduction">Toxin Production</a></li>
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 +
<b>
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<span style="color:green">
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<li>Resources</li>
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</span>
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</b>
 +
 
 +
<li><a href="https://2009.igem.org/Team:HKUST/Lab Notebook">Lab Notebook</a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/Parts">Parts Submitted </a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/Protocols">Protocol List</a></li>
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<li><a href="https://2009.igem.org/Team:HKUST/Resourses">Other Resources</a></li>
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</ul>
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</div>
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<div id="menubottom">
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<ul>
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<li><a href="https://2009.igem.org/Team:Gallery">Gallery</a></li>
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<li><a href="https://2009.igem.org/Team:Biosafety">Biosafety</a></li>
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<li><a href="https://2009.igem.org/Team:Acknowledgement">Acknowledgement</a></li>
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</ul>
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</div>
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</div>
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<div id="rightxx">
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<div class="contentlist"> <h3>a</h3>
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</div>
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<div class="contentxx">
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<p>Yeast genomic DNA extraction</p>
 +
 
 +
<p>   Purpose: To extract yeast genomic DNA from yeast strain (strain YPH501, in our case).</p>
 +
   Materials: yeast strain, lysis buffer, phenol, chloroform, 3M NaAc, 100% ethanol, 70% ethanol, ddH2O, glass beads. <br> <br>
 +
 
 +
<p>Procedure: </p>
 +
   a.Add 200μL lysis buffer.<br>
 +
   b.Pick a whole patch of colony and add it in the tube.<br>
 +
   c.Add 100μL phenol, and mix it well.<br>
 +
   d.Add 100μL chloroform, and a few glass beads.<br>
 +
   e.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm). <br>
 +
   f.Put the supernatant (~160μL) to another tube. <br>
 +
   g.Add 200μL phenol/chloroform (1:1 in volume)<br>
 +
   h.Vortex for 4-5mins and centrifuge for 5mins (13,000 rpm). <br>
 +
   i.Put the supernatant (~120μL) to another tube. <br>
 +
   j.Add 0.1 volume NaAc(pH5.2), approximately 12μL.<br>
 +
   k.Add 2 volume of 100% ethanol, approximately 240μL.<br>
 +
   l.Store it at -20 °C for 20mins.<br>
 +
   m.Centrifuge for 10mins (13,000 rpm) and remove the supernatant.<br>
 +
   n.Add 2 volume of 70% ethanol.<br>
 +
   o.Centrifuge for 5mins (13,000 rpm) and remove the supernatant. <br>
 +
   p.Stand the tube without closing it for a while to let the ethanol evaporate.<br>
 +
   q.Add 20μL ddH2O to resuspend it.<br>
 +
   r.Store it at -20 °C.<br><br>
 +
 
 +
<p> Safety tips: </p>
 +
Phenol is a strong toxin, make sure to wear gloves and be careful with this chemical. <br><br>
 +
 
 +
<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>
 +
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Transformation_to_yeast">Transformation to
 +
yeast</a></li>
 +
<li><a href="https://2009.igem.org/Team:HKUST/Protocols/Cell_lysis"> Cell lysis</a></li>
 +
<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>
 +
 
 +
</ul>
 +
 +
</div>
 +
<div class="productxx">
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 +
<div class="clear"></div>
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</div>
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</div>
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<div class="clear"></div>
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</div>
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</div>
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<div id="footerxx">
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<div id="copyright">
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<span> iGEM 2009 <br /> </span>
 +
</div>
 +
<div id="payment"><img src="http://igem2009hkust.fileave.com/wiki/template/12092009/images/HKUSTLogo.jpg" alt="HKUST" /></div>
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</div>
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<div id="footerend"></div>
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</div>
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</bodyxx>
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</html>

Latest revision as of 09:57, 20 October 2009

Salt and Soap template

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.

HKUST

Retrieved from "http://2009.igem.org/Team:HKUST/Protocols/Yeast_genomic_DNA_extraction"