Team:UNITN-Trento/Notebook/Labposts/07/19

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{
{
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"date" : "2013-07-22",
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"date" : "2013-07-10",
"author" : "fabio",
"author" : "fabio",
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"title" : " blue and red light sensors!!",
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"title" : " ripenator v1.0 ",
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"content" : "<html> let’s get ready to screen!! This time I amplified my parts by pcr them out with OneTaq polymerase!! I started from the blue promoter (number A from the firt  set). Pcr sample composition: <br>5X Onetaq standard reaction buffer  10 ul <br>10 mM dNTPs  1 ul<br>Forward primer 1ul<br>Reverse primer 1 ul<br>One Taq 0.25<br>Phusion 0.3<br>Template 0.5<br>Water up to 50 ul<br>Pcr program values: <br>Initial digestion: 94° 2min <br>Digestion: 94 30 sec<br>Annealing: 60 1 min<br>Annealing: 68° 15 sec<br>Final extention: 68° 5 min<br>(30 cycles from step 3 to step 5) <br>The screening was successful : <br> </html>{{:Team:UNITN-Trento/Templates/Styles/Spoiler|gel|<html><center><img src=\"https://static.igem.org/mediawiki/2013/0/06/Tn-2013_C.jpg\" width=\"450px\" /></center></html>}}<html>Finally I purified the PCR and abtained a 209,8 ng/ul yieald.Meanwhile I started thinking about the ligation of the two blue parts: the idea is to use k592016 as plasmid and k592006 as an insert to put downstream . So I had to digest o/n  3 ug of 016(from the first set)  with Spel and Pstl, and 006( the purified Pcr)  with Xbal and Pstl. At the same time I digested R0010 with Ecorl and Spel, cause I’ll need to put it on top of everything once I have my biobrick.</html>",
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"content" : "<html> this morning, Bruno is not available, so I will take care of the ripenator. I have to replace all the cultures with new LB and new bacteria : I’m gonna put 300 ml of LB in each beutas, then connect immediately a beuta with LB and 3ml of not-induced bacteria to the control banana!! After that, I will dissolve 78 ul of MESA in the second beuta and connect to the blocked banana! As far as the ethylene sample is concerned, I have to put 3 ml of bacteria in the last beuta, wait for the OD lecture to reach the value of 0,5 and then induce the culture with Arabinose to produce Ethylene! Finally I can connect it with the mature banana. </html>",
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"tags" : " YF1_FixJ - FixK2- ho1_PcyA_cph8-OmpR"
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"tags" : " ripenator-mesa-ethylene "
}
}

Revision as of 08:26, 3 October 2013

{ "date" : "2013-07-10", "author" : "fabio", "title" : " ripenator v1.0 ", "content" : " this morning, Bruno is not available, so I will take care of the ripenator. I have to replace all the cultures with new LB and new bacteria : I’m gonna put 300 ml of LB in each beutas, then connect immediately a beuta with LB and 3ml of not-induced bacteria to the control banana!! After that, I will dissolve 78 ul of MESA in the second beuta and connect to the blocked banana! As far as the ethylene sample is concerned, I have to put 3 ml of bacteria in the last beuta, wait for the OD lecture to reach the value of 0,5 and then induce the culture with Arabinose to produce Ethylene! Finally I can connect it with the mature banana. ", "tags" : " ripenator-mesa-ethylene " }

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