Difference between revisions of "Haynes Lab:Notebook/Protein-DNA fusions in living microbes/2013/10/09"

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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span>
 
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|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
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used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf
 
used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf
  
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Made SOC media:
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LB miler broth + tryptone + 20 mM MgSO4 and 2.5 mM KCl (http://openwetware.org/wiki/SOB)
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Grew up 4 colonies of BL21(DE3) in Kan media, used 3 of them (1, 2, 4) to induce expression of Topo per protocol: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf
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Ran PCR using 1-omega F TSE and alpha-4 R SP and PSV plasmid template. PCR product has 3 topo cut sites.
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Ran gel of wheat-germ topo +RFP pSB1C3 plasmid. From left to right: 2uL, 1.5uL, 1uL, 0.5uL, blank glycerol stock (15/30 uL in each well)
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Made competent cell stock of BL21(DE3) from cells given by Kylie
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Used Bug buster on induced samples (1, 2, 4) per protocol: http://www.ebiotrade.com/buyf/productsf/novagen/70584-000.pdf
  
 
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Latest revision as of 22:36, 26 September 2017

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Entry title

Performed successful tranformation with Kylie's BL21(DE3) used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf

Made SOC media: LB miler broth + tryptone + 20 mM MgSO4 and 2.5 mM KCl (http://openwetware.org/wiki/SOB)

Grew up 4 colonies of BL21(DE3) in Kan media, used 3 of them (1, 2, 4) to induce expression of Topo per protocol: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf

Ran PCR using 1-omega F TSE and alpha-4 R SP and PSV plasmid template. PCR product has 3 topo cut sites.

Ran gel of wheat-germ topo +RFP pSB1C3 plasmid. From left to right: 2uL, 1.5uL, 1uL, 0.5uL, blank glycerol stock (15/30 uL in each well)

Made competent cell stock of BL21(DE3) from cells given by Kylie

Used Bug buster on induced samples (1, 2, 4) per protocol: http://www.ebiotrade.com/buyf/productsf/novagen/70584-000.pdf