Team:TU Delft/28 July 2010 content

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=Lab work=
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==Hydrocarbon Sensing==
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[https://2010.igem.org/Team:TU_Delft#page=pages/blog&blog=27_July_2010 Yesterday's] transformations of the Biobricks AlkS-B0015 and AlkS-E0422 were checked with scPCR.
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The resulting gel is shown below.
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The colony #1 of AlkS-B015 and colony #2 AlkS-E0422 were used for plasmid isolation.
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==Alkane degradation==
==Alkane degradation==
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[[Image:TUDelft_20100728_scPCR1.png|400px|thumb|left|1% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker]]
[[Image:TUDelft_20100728_scPCR1.png|400px|thumb|left|1% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker]]
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'''Lane Description part 1'''
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Lane Description part 1
{| 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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[[Image:TUDelft_20100728_scPCR2b.png|400px|thumb|left|% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker]]
[[Image:TUDelft_20100728_scPCR2b.png|400px|thumb|left|% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker]]
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'''Lane Description part 2'''
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Lane Description part 2
{| 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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Latest revision as of 08:43, 13 October 2010

Lab work

Hydrocarbon Sensing

Yesterday's transformations of the Biobricks AlkS-B0015 and AlkS-E0422 were checked with scPCR.

The resulting gel is shown below.

The colony #1 of AlkS-B015 and colony #2 AlkS-E0422 were used for plasmid isolation.

Alkane degradation

The plates containing yesterday's ligations contained colonies, to check whether they really contain the desired BioBrick a colony PCR was done, and the used colonies were grown in liquid LB medium over night. The results from the PCR were analysed on a 1% agarose gel.

1% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker

Lane Description part 1

# Description Expected lenght (bp) Primers Status
1 SmartLadder n/a n/a n/a
2 J61100 + rubA3 473 G00100 + G00101 ✓
3 J61100 + rubA3 473 G00100 + G00101 ✓
4 J61100 + rubA3 473 G00100 + G00101 ✗
5 J61100 + rubA3 473 G00100 + G00101 ✓
6 J61100 + rubA3 473 G00100 + G00101 ✗
7 J61100 + rubA4 482 G00100 + G00101 ✓
8 J61100 + rubA4 482 G00100 + G00101 ✓
9 J61100 + rubA4 482 G00100 + G00101 ✓
10 J61100 + rubA4 482 G00100 + G00101 ✓
11 J61100 + rubA4 482 G00100 + G00101 ✓
12 J61100 + rubR 1505 G00100 + G00101 ✓
13 J61100 + rubR 1505 G00100 + G00101 ✓
14 J61100 + rubR 1505 G00100 + G00101 ✓
15 J61100 + rubR 1505 G00100 + G00101 ✓
16 J61100 + rubR 1505 G00100 + G00101 ✓
17 SmartLadder n/a n/a n/a
% agarose of colony PCR part 1. Gel runned at 100V for 1 hour. Of all samples 10 μL + 2 μL loadingbuffer was loaded. 5 μL was loaded of marker

Lane Description part 2

# Description Expected lenght (bp) Status Remarks
1 SmartLadder n/a n/a n/a
2 J61100 + ladA 1625 G00100 + G00101 ✓
3 J61100 + ladA 1625 G00100 + G00101 ✓
4 J61100 + ladA 1625 G00100 + G00101 ✓
5 J61100 + ladA 1625 G00100 + G00101 ✓
6 J61100 + ladA 1625 G00100 + G00101 ✗
7 J61101 + ADH 1052 G00100 + G00101 ✓
8 J61101 + ADH 1052 G00100 + G00101 ✓
9 J61101 + ADH 1052 G00100 + G00101 ✗
10 J61101 + ADH 1052 G00100 + G00101 ✓
11 J61101 + ADH 1052 G00100 + G00101 ✗
12 J61107 + ALDH 1796 G00100 + G00101 ✗
13 J61107 + ALDH 1796 G00100 + G00101 ✓
14 J61107 + ALDH 1796 G00100 + G00101 ✗
15 J61107 + ALDH 1796 G00100 + G00101 ✗
16 J61107 + ALDH 1796 G00100 + G00101 ✗
17 J61100 (uncut) G00100 + G00101 ✓
18 J61101 (uncut) G00100 + G00101 ✓
19 J61107 (uncut) G00100 + G00101 ✓
20 SmartLadder n/a n/a

From the scPCR it can be concluded that from all the colonies PCRed (per BioBrick), at least one was positive! Next week the next (last) step in making the BioBricks for alkane degradation can be be done, so hopefully the week after that we can start characterizing.

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