Team:TU Delft/28 July 2010 content

From 2010.igem.org

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.