Team:IPN-UNAM-Mexico/Notebook

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{{Template:IPN-UNAM-Mexico}}
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As summer proyect our experimental work began at 07 - july - 2009 using 2008 bioparts catalog, unfurtunly it dosen’t work properly, we had to much troubles to take out DNA and transform, E. Coli for this we had to delay and request 2009 catalog.
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=NOTEBOOK=
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Our proyect uses 29 bioparts, 12 ligations and diferent strategys to make it functional as follow:
 
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<h1> [[Image:Month-icon.png | 50px]] June</h1>
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 +
As summer proyect our experimental work began at july the 7th 2009 using 2008 bioparts catalog, unfortunately it didn't work properly, so we had a lot of trouble to take out DNA and transform into ''E. coli''. For this reason we had to delay lab work and request the 2009 catalog.
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<h1>[[Image:Month-icon.png | 50px]] July </h1>
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<h1>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Protocols|Protocols]]</h1>
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'''''07-July-2009'''''
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<h2>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Notebook/July|July]]</h2>
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<h2>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Notebook/August|August]]</h2>
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<h2>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Notebook/September|September]]</h2>
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<h2>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Notebook/October|October]]</h2>
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We digest the follow biobriks with ECORI (E), SPEI (S), and XBAI (X).
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<h1>[[Image:Month-icon.png | 50px]][[Team:IPN-UNAM-Mexico/Safety|Safety]]</h1>
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Our synthetic construction consists of 29 bioparts, which needed 12 ligations and different strategies to make it functional as is  described below:
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{| border="0.5"
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! Biobrick
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! Enzime restriction
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-
|-
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|R0079
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|E, S
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-
|-
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-
|F1610
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-
|E, X
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-
|-
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-
|B0034
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|E, S
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-
|-
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-
|C0078
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|E, X
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-
|-
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|C0079
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|E, X
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-
|-
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-
|B0015
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|E, X
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|}
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We propose this restrictions for standar assembly for the digest we use a 17 hours incubation at 37º camera.
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Digest Mix:
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{| border="0.5"
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! ERCO RI - SPEI
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! Per reaction
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|-
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|Plasmidic DNA
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|align="right" | 3μl
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-
|-
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|Enzima ECORI
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|align="right" | 2μl
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-
|-
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|Enzima SPEI
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|align="right" | 2μl
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|-
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|Buffer NBE
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|align="right" | 2μl
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-
|-
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|BSA
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|align="right" | 0.5μl
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|-
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|H2O
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|align="right" | 10.5μl
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|-
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!Total
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|align="right" | 20μl
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|}
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{| border="0.5"
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! ECORI - XBAI
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!Per reaction
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|-
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|Plasmidic DNA
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|align="right" | 3μl
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|-
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|Enzima ECORI
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|align="right" | 2μl
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|-
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|Enzima XBAI
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|align="right" | 2μl
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|-
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|Buffer NBE
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|align="right" | 2μl
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|-
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|BSA
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|align="right" | 0.5μl
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|-
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|H20
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|align="right" | 10.5μl
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|-
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!Total
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|align="right" | 20μl
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|}
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'''''08-July-2009'''''
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We carried out the restrictions on a 10 wells agarosa gel 40 min.
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3μl DNA  2μl Buffer.
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We have not  DNA on the Gels maybe the DNA volume is so low.  We try again with
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20μl DNA  3μl Buffer
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We run on  only Plasmidic DNA  but we have not DNA.
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'''''09-July-2009'''''
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We start from cero, and take off DNA from the folder plateing R0079, C0178, C0179, C0060, B0034, I739001 biobriks,
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We left 16 hours in 37º camera.
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'''''10-July-2009'''''
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We don’t have transform cells in this point we have to delay and request 2009 catalog.
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'''''28-July-2009'''''
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With the 2009 catalog we retake the work and take on the biobriks we need. First of all we did a test to be sure that the biobriks worked properly for this we take a biobrik with RFP reporter (BBa_I3522) and plate, we incubate 17 hours in 37º camera.
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'''''29-July-2009'''''
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The transformation was successful we are sure the biobricks and method works and we proceed to take off our biobricks as next chart.
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{| border="0.5"
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<center>
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{| border="2"
!Number
!Number
!Biobrick
!Biobrick
 +
!Restriction
|-
|-
|2
|2
|BBa_R0079
|BBa_R0079
 +
|S & P
|-
|-
|3
|3
|BBa_F1610
|BBa_F1610
 +
|X & P
|-
|-
|4
|4
|BBa_K091146
|BBa_K091146
 +
|S & P
|-
|-
|5
|5
|BBa_K093005
|BBa_K093005
 +
|X &
|-
|-
|6
|6
|BBa_K081016
|BBa_K081016
 +
|X & P
|-
|-
|7
|7
|BBa_EC840
|BBa_EC840
 +
|X & P
|-
|-
|8
|8
|BBa_K081009
|BBa_K081009
 +
|X & P
|-
|-
|9
|9
|BBa_R0051
|BBa_R0051
 +
|S &P
|-
|-
|10
|10
|BBa_J06800
|BBa_J06800
 +
|X & P
|-
|-
|11
|11
|BBa_Q04121
|BBa_Q04121
 +
|X & P
|-
|-
|12
|12
|BBa_B0034
|BBa_B0034
 +
|S & P
|-
|-
|13
|13
|BBa_C0079
|BBa_C0079
 +
|E & S
|-
|-
|14
|14
|BBa_C0179
|BBa_C0179
 +
|E & S
|-
|-
|15
|15
|BBa_B0015
|BBa_B0015
 +
|E & X
|-
|-
|16
|16
|BBa_K081018
|BBa_K081018
 +
|E & S
|-
|-
|17
|17
|BBa_K116640
|BBa_K116640
 +
|X & P
 +
|-
 +
|23
 +
|BBa_S03154
 +
|X & P
 +
|-
 +
|24
 +
|BBa_k145201
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|E & S
|}
|}
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</center>
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For transformation we use the same method as follow.
 
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Elusion: 3μl DNA from the well, and get in Eppendorf 1.5 ml with competent cells. We use electoporation shock to transform and recovery in  LB ampr  1 hour and a half  then we plate on petri dishes and get  incubate 17 hours.
 
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'''''31-July-2009'''''
 
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We plated  Top 10 biobricks F1610, K091146, K081016, K081009, R0051, K081018 and get incube 17 hours
 
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<h1>[[Image:Month-icon.png | 50px]] August </h1>
 
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<h1>[[Image:Month-icon.png | 50px]] September </h1>
 
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<h1>[[Image:Month-icon.png | 50px]] October </h1>
 
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<center>
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{| border="2"
 +
! Ligation
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! Nickname
 +
! Biobrick
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|-
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|4-23
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|Ciencias I
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|BBa_K226005
 +
|-
 +
|13-15
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|Ciencias II
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|BBa_K266002
 +
|-
 +
|Ciencias I-7
 +
|Ciencias III
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|BBa_K266006
 +
|-
 +
|24-17
 +
|Ciencias IV
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|BBa_K266001
 +
|-
 +
|2-3
 +
|Ciencias V
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|BBa_K266000
 +
|-
 +
|J231000-11
 +
|Ciencias VI
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|BBa_K266008
 +
|-
 +
|Amplicon - Ciencias II
 +
|Ciencias VII
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|BBa_K266003
 +
|-
 +
|Ciencias VI - 12
 +
|Ciencias VIII
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|BBa_K266009
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|-
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|Ciencias III - 19
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|Ciencias IX
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|BBa_K266006
 +
|-
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|J23100 – Ciencias VII
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|Ciencias X
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|BBa_K266004
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|-
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|Ciencias III - Ciencias V
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|Ciencias XI
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|BBa_K266007
 +
|-
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|Ciencias X - Ciencias IV
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|Ciencias XII
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|BBa_K266010
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|}</center>
{{Template:IPN-UNAM-Mexico-footer}}
{{Template:IPN-UNAM-Mexico-footer}}

Latest revision as of 01:29, 22 October 2009


BannerUNAM.jpg

NOTEBOOK

As summer proyect our experimental work began at july the 7th 2009 using 2008 bioparts catalog, unfortunately it didn't work properly, so we had a lot of trouble to take out DNA and transform into E. coli. For this reason we had to delay lab work and request the 2009 catalog.

Month-icon.pngProtocols

Month-icon.pngJuly

Month-icon.pngAugust

Month-icon.pngSeptember

Month-icon.pngOctober

Month-icon.pngSafety

Our synthetic construction consists of 29 bioparts, which needed 12 ligations and different strategies to make it functional as is described below:


Number Biobrick Restriction
2 BBa_R0079 S & P
3 BBa_F1610 X & P
4 BBa_K091146 S & P
5 BBa_K093005 X &
6 BBa_K081016 X & P
7 BBa_EC840 X & P
8 BBa_K081009 X & P
9 BBa_R0051 S &P
10 BBa_J06800 X & P
11 BBa_Q04121 X & P
12 BBa_B0034 S & P
13 BBa_C0079 E & S
14 BBa_C0179 E & S
15 BBa_B0015 E & X
16 BBa_K081018 E & S
17 BBa_K116640 X & P
23 BBa_S03154 X & P
24 BBa_k145201 E & S


Ligation Nickname Biobrick
4-23 Ciencias I BBa_K226005
13-15 Ciencias II BBa_K266002
Ciencias I-7 Ciencias III BBa_K266006
24-17 Ciencias IV BBa_K266001
2-3 Ciencias V BBa_K266000
J231000-11 Ciencias VI BBa_K266008
Amplicon - Ciencias II Ciencias VII BBa_K266003
Ciencias VI - 12 Ciencias VIII BBa_K266009
Ciencias III - 19 Ciencias IX BBa_K266006
J23100 – Ciencias VII Ciencias X BBa_K266004
Ciencias III - Ciencias V Ciencias XI BBa_K266007
Ciencias X - Ciencias IV Ciencias XII BBa_K266010

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