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454-pAd5.CMV-cre-iRFP - Revision history
2026-10-18T12:08:06Z
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Yingying.lu at 19:13, 4 January 2012
2012-01-04T19:13:33Z
<p></p>
<p><b>New page</b></p><div>Cloning of pAd5-CMV-cre-iRFP <br />
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Purpose: to clone pAd5-CMV-iRFP by gene synthesis&nbsp; to 418 pAd5.CMV-iRFP <br />
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Date: 2011-11-18 <br />
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Notebook:Lu yingying <br />
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Page: <br />
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Clone ID:454 <br />
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Experimenters: lu yingying<br>Results<br> <br />
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[[Image:454 PCR 11-18-2011.JPG]]<br> <br />
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PCR Cre with Cre-EcoR1-F + Cre-HindIII-BglII-R, using pTriEx-HTNCre(298) as template<br> <br />
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[[Image:454 digest insert+vector11-18-2011.JPG]]<br> <br />
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PCR product digest with EcoR1 + Bgl11,&nbsp; <br />
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pAd5.CMV-iRFP 418 vector digest with EcoR1 + BamH1 <br />
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band 1 show insert,band 2 show pAd5.CMV-iRFP 418 vector<br> <br />
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[[Image:454 digest detect11-21-2011.JPG]] <br />
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digest test:EcoR1+Xho1 <br />
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<br> <br />
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Date: 2011-11-22 <br />
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Notebook:Lu yingying <br />
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Page: <br />
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Clone ID:454 <br />
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Experimenters: lu yingying<br>Results<br> <br />
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[[Image:454 clone 11-22-2011.JPG]]<br> <br />
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&nbsp;PCR product digest with EcoR1 + Bgl11, pAd5.CMV-iRFP 418 vector digest with EcoR1 + BamH1 <br> <br />
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insert and vector <br />
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[[Image:454 digest detect11-27-2011.JPG]]<br> <br />
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digest with EcoR1 +Xho1<br> <br />
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<br> <br />
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== '''Cloning of pAd5.CMV-cre-iRFP''' ==<br />
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'''Purpose:''' Cloning of pAd5.CMV-cre-iRFP <br />
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'''Date: '''2011-12-14-- <br />
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'''Notebook:''' <br />
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'''Page:''' <br />
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'''Clone ID:454''' <br />
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'''Experimenters:''' yingying <br />
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== '''Results''' ==<br />
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=== Subcloning ===<br />
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Date: 2011-12-14 <br />
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[[Image:414,454 Clone 12-14-2011.JPG]]<br> <br />
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{| cellspacing="1" cellpadding="1" border="1" style="width: 288px; height: 80px;"<br />
|-<br />
| 454<br> <br />
| band1<br> <br />
| band2<br><br />
|-<br />
| <br> <br />
| PCR product<br> <br />
| pAd5.CMV-iRFP 418<br><br />
|-<br />
| digest&nbsp; <br> <br />
| EcoR1 + Bgl11<br> <br />
| EcoR1 + BamH1 <br><br />
|}<br />
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<br> <br />
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<br> <br />
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=== Colony Screening ===<br />
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Date: 2011-12-16<br> <br />
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Make miniprep and digest with EcoR1 +Xho1<br> <br />
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[[Image:414+454 digest detect 12-16-2011.JPG]] <br />
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&nbsp;<br> <br />
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Lane 1: may right ,need to sequence<br><br><br />
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Transfect into 293T cells ,then read fluorescence :Ex675nm,Em740nm<br />
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{| cellspacing="1" cellpadding="1" border="1" style="width: 345px; height: 126px;"<br />
|-<br />
| <br><br />
| Temperature(°C)<br><br />
| blank<br><br />
| blank<br><br />
| 454<br><br />
| 454<br><br />
|-<br />
| read14<br><br />
| 24.7<br><br />
| 0.501<br><br />
| 0.483<br><br />
| 1.065<br><br />
| 4.195<br><br />
|-<br />
| read30<br><br />
| 24.9<br><br />
| 1.743<br><br />
| 1.322<br><br />
| 2.207 <br><br />
| 2.343<br><br />
|-<br />
| read54<br><br />
| 24.8<br><br />
| 0.486<br><br />
| 0.899<br><br />
| 0.958<br><br />
| 0.898<br><br />
|}<br />
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<br><br> <br> <br />
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== '''Result intepretation/Summary''' ==<br />
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*Sequencing Result:<br><br />
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== '''Conclusion''' ==<br />
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Restriction digestion <br> <br> <br />
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=== ===<br />
<br />
<br> --[[User:Yingying.lu|Yingying.lu]] 08:54, 19 December 2011 (EST) <br />
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[[Category:Cloning]]</div>
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