Team:UNITN-Trento/Notebook/Labposts/06/57

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(Created page with "{ "date" : "2013-06-29", "author" : "emil", "title" : "Purification and screening of the GFPs E0840 and E0240", "content" : "<html> I did the miniprep of the 8 inocula(2 GFP...")
 
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{
{
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"date" : "2013-06-29",
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"date" : "2013-06-25",
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"author" : "emil",
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"author" : "caterina-michele",
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"title" : "Purification and screening of the GFPs E0840 and E0240",
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"title" : "The final device: BBa_J45320(placI and PchBA)+ BBa_K1065101(araC-pBAD promoter and BMST1) in pSB1C3",
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"content" : "<html> I did the miniprep of  the 8 inocula(2 GFP constructs) following the <a href=\"http://ita.promega.com/~/media/Files/Resources/ProtCards/Wizard%20Plus%20SV%20Minipreps%20DNA%20Purification%20System%20Quick%20Protocol.pdf\">vacuum protocol</a> then I have quantified the products with the following results:</html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Quantification|<html><center><table class=\"tn-sp-table\"><tr><th>Sample</th><th>Quantification</th></tr><tr><td>BBa_E0840/1</td><td> 484.6ng/&micro;l</td></tr><tr><td>BBa_E0840/2</td><td> 376ng/&micro;l</td></tr><tr><td>BBa_E0840/3</td><td> 481.8ng/&micro;l</td></tr><tr><td>BBa_E0840/4</td><td> 475ng/&micro;l</td></tr><tr><th>BBa_E0240/1</th><th> 361ng/&micro;l</th></tr><tr><td>BBa_E0240/2</td><td> 454.4ng/&micro;l</td></tr><tr><td>BBa_E0240/3</td><td> 368ng/&micro;l</td></tr><tr><td>BBa_E0240/4</td><td> 365.8ng/&micro;l</td></tr></table></center></html>}}<html>Then I did the screening of the samples E0840/1,E0840/3,E0240/2,E0240/4. I digested 800 ng of each sample with EcoR1-HF and Pst1-HF following the <a href=\"https://2013.igem.org/Team:UNITN-Trento/Protocols#Digestion\">digestion protocol for screening</a>. After 40 minutes I loaded the samples on a gel with the following results:</html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Gel order|<html><center><table class=\"tn-sp-table\"><tr><th>Sample</th><th>Well</th></tr><tr><td>Ladder 1kb Fermentas</td><td>1</td></tr><tr><td>BBa_E0840/1</td><td>2</td></tr><tr><td>BBa_E0840/3</td><td>3</td></tr><tr><td>BBa_E0240/2</td><td>4</td></tr><tr><td>BBa_E0240/4</td><td>5</td></tr></table></center></html>}}{{:Team:UNITN-Trento/Templates/Styles/Spoiler|image|<html> <img src=\"https://static.igem.org/mediawiki/2013/4/4b/Tn-20130629-Et_GFP.jpg\" width=\"450px\" /></html>}}<html> We can see 2 bands for well: the upper is the backbone(pSB1A2 2079 bp), the lower is presumably the GFP(BBa_E0240 826 bp,BBa_E0840 828 bp)(pefix and suffix escluded ?).</html>",
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"content" : " We performed the digestion and the ligation on:1a)BBa_K1065101 with EcoR I and Xba I and1b)BBa_J45320 with EcoR I and Spt I to obtain the complete device BBa_K1065102(placI+PchBA+araC-pBAD promoter+BMST1 in pSB1C3)2a)pSB1C3 linearized with EcoR I and Pst I2b)BBa_J45320 with EcoR I and Pst Ito obtain the device BBa_K1065103 (placI+PchBA in pSB1C3).At the end of the day we did the trasformaion in NEB10&beta; cells. Waiting for the results...break a leg! ",
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"tags" : "E0840-E0240"
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"tags" : "PchBA-BMST1-araC-pBAD"
}
}

Latest revision as of 07:57, 3 October 2013

{ "date" : "2013-06-25", "author" : "caterina-michele", "title" : "The final device: BBa_J45320(placI and PchBA)+ BBa_K1065101(araC-pBAD promoter and BMST1) in pSB1C3", "content" : " We performed the digestion and the ligation on:1a)BBa_K1065101 with EcoR I and Xba I and1b)BBa_J45320 with EcoR I and Spt I to obtain the complete device BBa_K1065102(placI+PchBA+araC-pBAD promoter+BMST1 in pSB1C3)2a)pSB1C3 linearized with EcoR I and Pst I2b)BBa_J45320 with EcoR I and Pst Ito obtain the device BBa_K1065103 (placI+PchBA in pSB1C3).At the end of the day we did the trasformaion in NEB10β cells. Waiting for the results...break a leg! ", "tags" : "PchBA-BMST1-araC-pBAD" }

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