Editing 1062-pLVX-CAG-Dest-IRES-EGFP

Cloning of pLVX-CAG-Dest-IRES-EGFP(1062)<br>Purpose: to clone pLVX-CAG-Dest-IRES-EGFP<br>Date: 2012-4-19 to 2012-4-<br>Notebook: Xiaojing Wang<br>Page: 004228 

Clone ID: 1062<br>Experimenters: Xiaojing Wang 

Results 

PCR gateway cassette using GW-AgeI-F + GW-Flag-R-Xho1 and pVP16,446 as template<br> 

[[Image:PCR gateway cassett 4 17 2012.jpg]]<br> 

Cut PCR product with Age1 + Xho1<br>Cut vector 1061 pLVX-CAG-MCS-IRES-EGFP with Age1 + Xho1 <br> 

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ligate and transform ccdbSurvival<br> 

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5/1/2012<br> 

Method 2<br>PCR using ccdb-attR1-EcoR1-F &amp; ccdb-attR2-Xho1-R and pVP16,446 as template--reducted ccdb gateway<br> 

[[Image:1062 1063 PCR 5.1.2012.JPG|106x251px|1062 1063 PCR 5.1.2012.JPG]]<br> 

Digest with EcoR1 + Xho1, clone into EcoR1 + Xho1 digested pLVX-CAG-MCS-IRES-EGFP, 1061.<br> 

[[Image:1062 1063 P V digest 5.1.2012.JPG]]<br> 

Ligate and transform ccdb Survival<br> 

Select 4 clones to mini-prep<br>1062 plasmids are digested by EcoR1+Xho1<br> 

[[Image:1062 1063 digest 5.3.2012.JPG]] 

<br>pDonr221_EGFP(393) + 1062-1 make gateway by LR Clonase II. 

Transform 393+1062-1 by STBL3, Amp. 

Select 4 clones to mini-prep.<br> 

1062+393 plasmids are digested by EcoR1+Xho1<br> 

[[Image:1062+393 1063+393 digest 5.8.2012.JPG]]<br> 

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== '''Cloning of 1062-pLVX-CAG-Dest-IRES-EGFP'''  ==

'''Purpose:''' Cloning of 1062-pLVX-CAG-Dest-IRES-EGFP 

'''Date: '''2012-5-14--- 

'''Notebook:0''' 

'''Page:''' 

'''Clone ID:1062''' 

'''Experimenters:''' yingying 

== '''Results'''  ==

=== cloning  ===

[[Image:1063 PCR 5-14-2012.JPG]]<br> 

The same to 1063 PCR prodedure, 

PCR gateway cassette using GW-AgeI-F + GW-Flag-R-Xho1 and pVP16,446 as template<br> 

[[Image:537+1035+1062+1063clone 5-15-2012.JPG]]<br> 

{| cellspacing="1" cellpadding="1" border="1" style="width: 407px; height: 83px;"
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| band 5<br> 
| band 4<br>
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| PCR product<br> 
| 1061 pLVX-CAG-MCS-IRES-EGFP<br>
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| enzymes<br> 
| colspan="2" | &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; Age1 + Xho1<br>
|}

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=== Colony Screening  ===

Date: 2012-5-17 

Make miniprep and digest with EcoR1+Xho1<br> 

[[Image:1062+1063 digest detect 5-17-2012.JPG]] 

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c <br> 

== '''Result intepretation/Summary'''  ==

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== '''Conclusion'''  ==

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== '''Cloning of pLVX-CAG-Dest-IRES-EGFP'''  ==

'''Purpose:''' pLVX-CAG-Dest-IRES-EGFP 

'''Date: '''2013-3- 

'''Notebook:''' 

'''Page:''' 

'''Clone ID:1062''' 

'''Experimenters:''' Shanyi 

== '''Results'''  ==

=== cloning  ===

Date:2013-3-<br> 

PCR gateway cassette using GW-AgeI-F + GW-Flag-R-Xho1 and pVP16,446 as template, <br>

cut PCR product with Age1 + Xho1 <br>

Cut vector 1061 pLVX-CAG-MCS-IRES-EGFP with Age1 + Xho1 <br>

ligate and transform ccdbSurvival 

[[Image:1062_1063pcr.JPG‎]]<br>

digest&nbsp;: 

[[Image:Digest_for_1062.JPG‎]]<br> 

{| cellspacing="1" cellpadding="1" border="1" style="width: 298px; height: 82px;"
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| band 3,4<br>
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| &nbsp; 1061pLVX-CAG-MCS-IRES-EGFP <br> 
| PCR<br>
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| enzymes<br> 
| colspan="2" | &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; Nco1+ Xho1<br>
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=== Colony Screening  ===

Date: 2013 

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== '''Result intepretation/Summary'''  ==

*<br>

== '''Conclusion'''  ==

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[[Category:Cloning]]