Team:TU Delft/4 August 2010 content
From 2010.igem.org
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[[Image:TUDelft_20100804_PCR1.png|350px|thumb|left|PCR 1]] | [[Image:TUDelft_20100804_PCR1.png|350px|thumb|left|PCR 1]] | ||
- | + | '''Lane description''' | |
- | Lane description | + | |
{| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1" | {| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1" | ||
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|011A | |011A | ||
|G00100 + G00101 | |G00100 + G00101 | ||
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|none | |none | ||
|6,7,8,9,10 | |6,7,8,9,10 | ||
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|012K | |012K | ||
|G00100 + G00101 | |G00100 + G00101 | ||
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|3,5 | |3,5 | ||
|1,2,4 | |1,2,4 | ||
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[[Image:TUDelft_20100804_PCR2.png|350px|thumb|left|PCR 2]] | [[Image:TUDelft_20100804_PCR2.png|350px|thumb|left|PCR 2]] | ||
+ | '''Lane description''' | ||
+ | {| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1" | ||
+ | |'''#''' | ||
+ | |'''Description''' | ||
+ | |'''Primers''' | ||
+ | |'''Expected length (bp)''' | ||
+ | |'''✓''' | ||
+ | |'''✗''' | ||
+ | |- | ||
+ | |L | ||
+ | |SmartLadder | ||
+ | |n/a | ||
+ | |n/a | ||
+ | |n/a | ||
+ | |n/a | ||
+ | |- | ||
+ | |1-5 | ||
+ | |020K | ||
+ | |G00100 + G00101 | ||
+ | |2428 | ||
+ | |none | ||
+ | |1,2,3,4,5 | ||
+ | |- | ||
+ | |6-10 | ||
+ | |021A | ||
+ | |G00100 + G00101 | ||
+ | |2011 | ||
+ | |8,10 | ||
+ | |6,7,9 | ||
+ | |- | ||
+ | |11-15 | ||
+ | |021K | ||
+ | |G00100 + G00101 | ||
+ | |2011 | ||
+ | |11,12,13,15 | ||
+ | |14 | ||
+ | |- | ||
+ | |L | ||
+ | |SmartLadder | ||
+ | |n/a | ||
+ | |n/a | ||
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+ | |n/a | ||
+ | |} |
Revision as of 15:16, 5 August 2010
Alkane Sensing, Solvent Tolerance, Salt Tolerance, Emulsifier
by Pieter
We found it remarkable that the only positive samples we found so far were from the series without preamplifying the insert by PCR (e.g. the AlkS). This is why we decided to retry the insertion of bbc1, PhPFDalpha and PhPFDbeta in J61101, but this time without PCRing the insert.
Digestion
For the assembly of BBa_K398101, BBa_K398205, BBa_K398206, BBa_K398402 and BBa_K398403 a digestion was done:
# | Sample | Enzyme 1 | Enzyme 2 | Enzyme 3 | Buffer | BSA | Needed fragment |
1 | 1 μg bbc1 | XbaI | PstI | ScaI | 2 (Biolabs) | ✓ | ‘X – bbc1 - P’ |
2 | 1 μg PhPFDalpha | XbaI | PstI | ScaI | 2 (Biolabs) | ✓ | ‘X – PhPFDalpha - P’ |
3 | 1 μg PhPFDbeta | XbaI | PstI | ScaI | 2 (Biolabs) | ✓ | ‘X – PhPFDbeta - P’ |
4 | 1 μg J61101 | SpeI | PstI | 2 (Biolabs) | ✓ | ‘S – J61101 - P’ | |
5 | 1 μg OprG-B0015 | XbaI | PstI | 2 (Biolabs) | ✓ | ‘X – OprG-B0015 - P’ | |
6 | 1 μg AlnA-B0015 | XbaI | PstI | 2 (Biolabs) | ✓ | ‘X – AlnA-B0015 - P’ | |
7 | 1 μg R0011-B0032 (sample 9) | SpeI | PstI | 2 (Biolabs) | ✓ | ‘S – R0011-B0032 - P’ | |
8 | 1 μg R0011-B0032 (sample 10) | SpeI | PstI | 2 (Biolabs) | ✓ | ‘S – R0011-B0032 - P’ |
Alkane degradation
Today we started with a PCR of a number colonies from yesterday's transformants. These had all given colonies on the plates, so hopefully they also contain the BioBricks! Five colonies from each plate were taken for the colony PCR, and after the PCR the products were analyzed on two gels.
Lane description
# | Description | Primers | Expected length (bp) | ✓ | ✗ |
L | SmartLadder | n/a | n/a | n/a | n/a |
1-5 | 007A | G00100 + G00101 | 1538 | 1,2,4 | 3,5 |
6-10 | 011A | G00100 + G00101 | 1816 | none | 6,7,8,9,10 |
11-15 | 012K | G00100 + G00101 | 1720 | 3,5 | 1,2,4 |
L | SmartLadder | n/a | n/a | n/a | n/a |
Lane description
# | Description | Primers | Expected length (bp) | ✓ | ✗ |
L | SmartLadder | n/a | n/a | n/a | n/a |
1-5 | 020K | G00100 + G00101 | 2428 | none | 1,2,3,4,5 |
6-10 | 021A | G00100 + G00101 | 2011 | 8,10 | 6,7,9 |
11-15 | 021K | G00100 + G00101 | 2011 | 11,12,13,15 | 14 |
L | SmartLadder | n/a | n/a | n/a | n/a |