Team:UNITN-Trento/Notebook/Labposts/06/49

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Latest revision as of 07:54, 3 October 2013

{ "date" : "2013-06-22", "author" : "bruno", "title" : "Minipreps and screening of Promoters", "content" : "From inocula of the day before I made the minipreps by the followingprotocol.

Quantification results

Sample	ng/ul
K090501	117,1
K090504_a	115,3
K090504_b	101,3
K823000_a	156,0
K823000_b	171,1
K823000	115,5

I made the screening test. To do that we digested 1500 ng of DNA with EcorI and PstI following the screening digestion protocol.
I used a 1.5% concentration of agarose to create a gel and I used Ethidium bromide instead the normal EuroSafe. In addition I have not used a normal Dye with the 20ul of sample loaded but I used 4ul of 30% glycerol. ", "tags" : "lacOid-PliaG-Pgram+" }

@@ Line 1: / Line 1: @@
 {
-	"date" : "2013-06-25",
+	"date" : "2013-06-22",
-	"author" : "emil",
+	"author" : "bruno",
-	"title" : "Purification and digestion of R0010",
+	"title" : "Minipreps and screening of Promoters",
-	"content" : "<html>I purified, and then quantified, 3 out of 5 of the previous inocula following the <a href=\"https://2013.igem.org/Team:UNITN-Trento/Protocols#miniprep\">miniprep protocol with vacuum</a>.</html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Quantification results|<html><center><table class=\"tn-sp-table\"><tr><th>Sample</th><th>Quantity</th></tr><tr><td>1</td><td>396.1ng/&micro;l</td></tr><tr><td>2</td><td>523.1ng/&micro;l</td></tr><tr><td>3</td><td>171ng/&micro;l</td></tr></table></center></html>}}<html>Afterwards, I digested 800ng of each sample  with Spe1-HF and Pst1-HF following the <a href=\"https://2013.igem.org/Team:UNITN-Trento/Protocols#Digestion\">digestion protocol for screening</a>. Finally I ran the products on a gel (1,5% agarose, due to the short length of the insert: only 200 bp + prefix and suffix). I loaded 12&micro;l of a ladder 100bp prepared as follows:</html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Ladder|<html><center><table class=\"tn-sp-table\"><tr><td style=\"border: none;\"></td><th>Quantity</th></tr><tr><td>Water</td><td>4&micro;l</td></tr><tr><td>Sharpmass Euroclone 100bp ladder </td><td>1&micro;l</td></tr><tr><td>glicerol solution(30%)</td><td>1&micro;l</td></tr></table></center></html>}}<html>This time, I loaded 12&micro;l (10&micro;l of sample and 2&micro;l of glicerol solution 30%) of each sample.</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>2&micro;l loading dye</td><td>1</td></tr><tr><td>Ladder</td><td>2</td></tr><tr><td>sample 1</td><td>3</td></tr><tr><td>sample 2</td><td>4</td></tr><tr><td>sample 3</td><td>5</td></tr><tr><td>sample 4(Bruno e Fabio)</td><td>6</td></tr><tr><td>sample 4(Bruno e Fabio)</td><td>6</td></tr><tr><td>Ladder 1000 kb</td><td>7</td></tr><tr><td>loading dye</td><td>8</td></tr></table></center></html>}}{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Gel Image|<html><img src=\"https://static.igem.org/mediawiki/2013/b/b4/Tn-20130625-EG_FB_20min.jpg\" width=\"450px\"></html>}}<html>We can see 2 bands: the highest is the backbone (pSB1A2: 2079bp) and the lower is pLac (nearly 243bp, located between the bands 300 and 200bp of the ladder).<br/>After the screening I digested 3&micro;g of the pSB1A2+R0010 (sample #2) with SpeI and PstI-HF o/n following the usual protocol.</html>",
+	"content" : "<html>From inocula of the day before I made the minipreps by the following<a href=\"https://2013.igem.org/Team:UNITN-Trento/Protocols#miniprep\">protocol</a>.</html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|Quantification results|<html><center><table class=\"tn-sp-table\"><tr><th>Sample</th><th>ng/ul</th></tr><tr><td>K090501</td><td>117,1</td></tr><tr><td>K090504_a</td><td>115,3</td></tr><tr><td>K090504_b</td><td>101,3</td></tr><tr><td>K823000_a</td><td>156,0</td></tr><tr><td>K823000_b</td><td>171,1</td></tr><tr><td>K823000</td><td>115,5</td></tr></table></center></html>}}<html>I made the screening test. To do that we digested 1500 ng of DNA with EcorI and PstI following <a href=\"https://2013.igem.org/Team:UNITN-Trento/Protocols#Digestion\">the screening digestion protocol</a>. <br>I used a 1.5% concentration of agarose to create a gel and I used Ethidium bromide instead the normal EuroSafe. In addition I have not used a normal Dye with the 20ul of sample loaded but I used 4ul of 30% glycerol. </html>",
-	"tags" : "Plac"
+	"tags" : "lacOid-PliaG-Pgram+"
 }