Team:Groningen/Notebook/Wetwork 23July2012

From 2012.igem.org

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Tom and Arjan started with gas chromatography mass spectrometry. Test sample: 5gr. fresh meat incubated @ 37C for 3 houres. For both column (HP5 and CP wax) three runs on each column with different incubation and syringe temperature (1: 80C, 100C 2: 100C, 120C 3: 120C, 140C)
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Nisa
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Plasmid isolation pSB1C3-alsT promoter + GFP
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<body><br><br><br>
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<p class="margin">
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Tom and Arjan started with gas chromatography mass spectrometry. Test sample: 5gr. fresh meat incubated @ 37C for 3 houres. For both column (HP5 and CP wax) three runs on each column with different incubation and syringe temperature (1: 80C, 100C 2: 100C, 120C 3: 120C, 140C)<br>
 +
<br>
 +
<br>
 +
Nisa<br>
 +
<br>
 +
Plasmid isolation pSB1C3-alsT promoter + GFP<br>
 +
<br>
 +
NanoDrop result (ng/ul):<br>
 +
A1= 102.9<br>
 +
A2= 96,6<br>
 +
B1= 118,1<br>
 +
B2= 113,7<br>
 +
C1= 122,8<br>
 +
C2= 88 Comment: gel picture OK!<br>
 +
D1= 124,9<br>
 +
D2= 93,3<br>
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<br>
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Result:<br>
 +
expected size: 3300bp for the pSB1C3 (2079bp)+ alsT (~500bp)+ GFP (800bp)<br>
 +
<br>
 +
From the gel picture, only plasmid from colony C2 (promoter alsT300) showed the expected size. The other plasmids seem to be empty, probably due to self ligation.<br>
 +
<br>
 +
Solution: Check plasmid from other white colony<br>
 +
<br>
 +
re do transformation. Increase the ratio of plasmid backbone: insert gene<br>
 +
<br>
 +
<br>
 +
Building backbone biobrick for B.subtilis<br>
 +
<br>
 +
plasmid pSac-Cm: integration plasmid, double cross over in SacA region<br>
 +
<br>
 +
add double terminator, BBa_B0015, and prefix-suffix: XbaI, SpeI, PstI<br>
 +
<br>
 +
<br>
 +
How to? <br>
 +
<br>
 +
Cut backbone with HindIII and EcoRI<br>
 +
<br>
 +
Cut BBa_B0015 with HindIII and EcoRI<br>
 +
<br>
 +
Now we have separate parts ready to use for ligation into our biobrick device<br>
 +
<br>
 +
<br>
 +
<A HREF="https://2012.igem.org/Team:Groningen/Notebook"><FONT COLOR=#ff6700>Back to notebook</FONT></A>
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<br><br></p></body></html>
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NanoDrop result (ng/ul):
+
{{Template:SponsorsGroningen2012}}
-
 
+
-
A1= 102.9
+
-
 
+
-
A2= 96,6
+
-
 
+
-
B1= 118,1
+
-
 
+
-
B2= 113,7
+
-
 
+
-
C1= 122,8
+
-
 
+
-
C2= 88 Comment: gel picture OK!
+
-
 
+
-
D1= 124,9
+
-
 
+
-
D2= 93,3
+
-
 
+
-
 
+
-
Result:
+
-
 
+
-
expected size: 3300bp for the pSB1C3 (2079bp)+ alsT (~500bp)+ GFP (800bp)
+
-
 
+
-
From the gel picture, only plasmid from colony C2 (promoter alsT300) showed the expected size. The other plasmids seem to be empty, probably due to self ligation.
+
-
 
+
-
Solution: Check plasmid from other white colony
+
-
 
+
-
re do transformation. Increase the ratio of plasmid backbone: insert gene
+
-
 
+
-
 
+
-
Building backbone biobrick for B.subtilis
+
-
 
+
-
plasmid pSac-Cm: integration plasmid, double cross over in SacA region
+
-
 
+
-
add double terminator, BBa_B0015, and prefix-suffix: XbaI, SpeI, PstI
+
-
 
+
-
 
+
-
How to?
+
-
 
+
-
Cut backbone with HindIII and EcoRI
+
-
 
+
-
Cut BBa_B0015 with HindIII and EcoRI
+
-
 
+
-
Now we have separate parts ready to use for ligation into our biobrick device
+

Latest revision as of 21:20, 25 September 2012







Tom and Arjan started with gas chromatography mass spectrometry. Test sample: 5gr. fresh meat incubated @ 37C for 3 houres. For both column (HP5 and CP wax) three runs on each column with different incubation and syringe temperature (1: 80C, 100C 2: 100C, 120C 3: 120C, 140C)


Nisa

Plasmid isolation pSB1C3-alsT promoter + GFP

NanoDrop result (ng/ul):
A1= 102.9
A2= 96,6
B1= 118,1
B2= 113,7
C1= 122,8
C2= 88 Comment: gel picture OK!
D1= 124,9
D2= 93,3

Result:
expected size: 3300bp for the pSB1C3 (2079bp)+ alsT (~500bp)+ GFP (800bp)

From the gel picture, only plasmid from colony C2 (promoter alsT300) showed the expected size. The other plasmids seem to be empty, probably due to self ligation.

Solution: Check plasmid from other white colony

re do transformation. Increase the ratio of plasmid backbone: insert gene


Building backbone biobrick for B.subtilis

plasmid pSac-Cm: integration plasmid, double cross over in SacA region

add double terminator, BBa_B0015, and prefix-suffix: XbaI, SpeI, PstI


How to?

Cut backbone with HindIII and EcoRI

Cut BBa_B0015 with HindIII and EcoRI

Now we have separate parts ready to use for ligation into our biobrick device


Back to notebook

Our sponsors:

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