Team:TU Delft/16 September 2010 content

From 2010.igem.org

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So the conclusion is that there is no difference between the control and the transformed cultures. So we just have to wait until we have the K12 strain.
So the conclusion is that there is no difference between the control and the transformed cultures. So we just have to wait until we have the K12 strain.
 +
 +
==Alkane degradation==
 +
009A and 010A were grown up overnight, and isolated using a mini-prep. The plasmids were digested and checked on a gel (cut and uncut). The gel looked good - no 'disappeared' plasmids as yesterday.
 +
 +
{| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1"
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|'''#'''
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|'''BioBrick'''
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|'''Cut with'''
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|'''Buffer'''
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|'''BSA'''
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|'''Expected Length (bp)'''
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|'''Correct?'''
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|-
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|0
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|Smartladder (5μl)
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|
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|
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|
 +
|
 +
|
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|-
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|1
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|009
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|uncut
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|
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|
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|
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|Yes
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|-
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|2
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|009
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|SpeI, PstI
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|Buffer 2 (NEB)
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|BSA (1x)
 +
|
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|Yes
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|-
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|3
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|010
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|uncut
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|
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|
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|
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|Yes
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|-
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|4
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|010
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|XbaI, PstI
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|Buffer 2 (NEB)
 +
|BSA (1x)
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|
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|Yes
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|-
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|5
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|018
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|uncut
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|
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|
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|
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|
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|-
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|6
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|018
 +
|EcoRI, SpeI
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|Buffer 2 (NEB)
 +
|BSA (1x)
 +
|
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|Yes
 +
|-
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|7
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|019
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|Uncut
 +
|
 +
|
 +
|
 +
|Yes
 +
|-
 +
|8
 +
|019
 +
|EcoRI, XbaI
 +
|Buffer 2 (NEB)
 +
|BSA (1x)
 +
|
 +
|Yes
 +
|-
 +
|9
 +
|001
 +
|uncut
 +
|
 +
|
 +
|
 +
|Yes
 +
|-
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|10
 +
|005
 +
|Uncut
 +
|
 +
|
 +
|
 +
|Yes
 +
|-
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|11
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|021AK
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|uncut
 +
|
 +
|
 +
|
 +
|Yes
 +
|-
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|}
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[[Image:TUDelft_20100916_digestioncheck.png|300px]]
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 +
The digestion products were ligated, and commercial TOP10 cells were transformed with the ligation mixtures.
 +
 +
{| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1"
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|'''BioBrick'''
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|'''Plasmid BB'''
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|'''Insert'''
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|'''Product'''
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|-
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|028A
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|009A
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|010A
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|J61100-rubA4-J61100-rubR
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|-
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|026A
 +
|018A
 +
|019A
 +
|J61101-ADH-J61107-ALDH
 +
|-
 +
|}

Latest revision as of 12:16, 1 October 2010

Emulsifier

Yesterdays cultures grown over night. Since Top10 cells activate Lac promoters by themselves, I did a quick and dirty experiment on those cultures.

1 ml culture was spin down for 2 min at max speed.

The supernatant was used in the emulsification assay.

# Description OD600 absorption (493 nm) absorption (493 nm) with Sudan II
1 Top 10 (control) 2.177 2.302
2 Top 10 with 206 1.703 1.862
3 Sup Top 10 (control) 0.013 0.341
4 Sup Top 10 with 206 0.015 0.349

So the conclusion is that there is no difference between the control and the transformed cultures. So we just have to wait until we have the K12 strain.

Alkane degradation

009A and 010A were grown up overnight, and isolated using a mini-prep. The plasmids were digested and checked on a gel (cut and uncut). The gel looked good - no 'disappeared' plasmids as yesterday.

# BioBrick Cut with Buffer BSA Expected Length (bp) Correct?
0 Smartladder (5μl)
1 009 uncut Yes
2 009 SpeI, PstI Buffer 2 (NEB) BSA (1x) Yes
3 010 uncut Yes
4 010 XbaI, PstI Buffer 2 (NEB) BSA (1x) Yes
5 018 uncut
6 018 EcoRI, SpeI Buffer 2 (NEB) BSA (1x) Yes
7 019 Uncut Yes
8 019 EcoRI, XbaI Buffer 2 (NEB) BSA (1x) Yes
9 001 uncut Yes
10 005 Uncut Yes
11 021AK uncut Yes

TUDelft 20100916 digestioncheck.png

The digestion products were ligated, and commercial TOP10 cells were transformed with the ligation mixtures.

BioBrick Plasmid BB Insert Product
028A 009A 010A J61100-rubA4-J61100-rubR
026A 018A 019A J61101-ADH-J61107-ALDH
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