Paul.xu at 19:32, 21 May 2012

2012-05-21T19:32:21Z

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Cloning of pLVX-CAG-Dest-IRES-mCherry(1063) Purpose: to clone pLVX-CAG-Dest-IRES-mCherry Date: 2012-4-12 to 2012-4- Notebook: Xiaojing Wang Page: 004224,004215 

Clone ID: 1063 Experimenters: Xiaojing Wang 

Results 

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

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

 

4/17/2012: 

Cut PCR product with Age1 + Xho1 Cut vector 1060 pLVX-CAG-MCS-CMV-mCherry with Age1 + Xho1

[[Image:1063 digestion 4 17 2012.jpg]] 

 

 ligate and transform ccdbSurvival 

Select 1 clone to do PCR screening 4/19/2012 

[[Image:1063 205 PCR Screening 4.19.2012.JPG]]

mini-prep, digest 4/20/2012

[[Image:1063 205 digest 4.20.2012.JPG]]

[[Image:1063 digest2 4.20.2012.JPG|83x240px|1063 digest2 4.20.2012.JPG]]

 

5/1/2012

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

[[Image:1062 1063 PCR 5.1.2012.JPG|93x220px|1062 1063 PCR 5.1.2012.JPG]] 

Digest with EcoR1 + Xho1, clone into EcoR1 + Xho1 digested pLVX-CAG-MCS-IRES-mCherry, 1060. 

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

Ligate and transform ccdb Survival 

Select 4 clones to mini-prep 1063 plasmids are digested by EcoR1+Xho1 

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

pDonr221_EGFP(393) + 1063-1 make gateway by LR Clonase II.

Transform 393+1063-1 by STBL3, Amp.

Select 1 clones to mini-prep.

1063+393 plasmid is digested by EcoR1+Xho1 

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

 

== '''Cloning of 1063-pLVX-CAG-Dest-IRES-mCherry''' ==

'''Purpose:''' Cloning of 1063-pLVX-CAG-Dest-IRES-mCherry

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

'''Notebook:0'''

'''Page:'''

'''Clone ID:1063'''

'''Experimenters:''' yingying

== '''Results''' ==

=== cloning ===

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

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

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

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| band 5 
| band 6 
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| PCR product 
| 1060,pLVX-CAG-MCS-CMV-mCherry 
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| enzymes 
| colspan="2" |            Age1 + Xho1 
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=== Colony Screening ===

Date: 2012-5-17

Make miniprep and digest with EcoR1+Xho1 

[[Image:1062+1063_digest_detect_5-17-2012.JPG]]

 

c 

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

*&lt;br&gt;

== '''Conclusion''' ==

&lt;br&gt;

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[[Category:Cloning]]

1063-pLVX-CAG-Dest-IRES-mCherry - Revision history

Paul.xu at 19:32, 21 May 2012