Team:TUDelft/30 July 2009

From 2009.igem.org

(Difference between revisions)
(→Calin)
(→Calin)
Line 98: Line 98:
Ran gel on backbones and oriT digest. Gel left for too long.
Ran gel on backbones and oriT digest. Gel left for too long.
 +
 +
[[Image:Gel30072009.png|550px]]
 +
 +
 +
{| border="1" align="center"
 +
| Well || Part || Plasmid Size
 +
|- align="center"
 +
| 1 || rbs-GFP-term I ||
 +
|- align="center"
 +
| 2 || rbs-GFP-term II ||
 +
|- align="center"
 +
| 3 || pSB1A3 I || 2832
 +
|- align="center"
 +
| 4 || pSB1A3 II || 2832
 +
|- align="center"
 +
| 5 || pSB1A3 III || 2832
 +
|- align="center"
 +
| 6 || pSB1A3 IV || 2832
 +
|- align="center"
 +
| 7 || pSB4A5 I || 4070
 +
|- align="center"
 +
| 8 || pSB4A5 II || 4070
 +
|- align="center"
 +
| 9 || pSB4A5 III || 4070
 +
|- align="center"
 +
| 10 || pSB4A5 IV || 4070
 +
|- align="center"
 +
| 11 || [http://www.eurogentec.com/EGT/Images/RESALES/Electrophoresis/Regular%20DNA%20Ladder/7-SmartLadder.jpg DNA Ladder] ||
 +
|- align="center"
 +
| 12 || pSB1C3 I || 2747
 +
|- align="center"
 +
| 13 || pSB1C3 II || 2747
 +
|- align="center"
 +
| 14 || pSB1C3 III || 2747
 +
|- align="center"
 +
| 15 || pSB1C3 IV || 2747
 +
|- align="center"
 +
| 16 || oriT E + P digest ||
 +
|- align="center"
 +
| 17 || oriT E + P digest ||
 +
|- align="center"
 +
| 18 || [http://www.eurogentec.com/EGT/Images/RESALES/Electrophoresis/Regular%20DNA%20Ladder/7-SmartLadder.jpg DNA Ladder] ||
 +
|}
===Tim Vos===
===Tim Vos===

Revision as of 21:38, 30 July 2009

Lab Notebook

July
MTWTFSS
    1 2 3 4 5
6 7 8 9 10 11 12
13 14 15 16 17 18 19
20 21 22 23 24 25 26
27 28 29 30 31
August
MTWTFSS
          1 2
3 4 5 6 7 8 9
10 11 12 13 14 15 16
17 18 19 20 21 22 23
24 25 26 27 28 29 30
31
September
MTWTFSS
  1 2 3 4 5 6
7 8 9 10 11 12 13
14 15 16 17 18 19 20
21 22 23 24 25 26 27
28 29 30
October
MTWTFSS
      1 2 3 4
5 6 7 8 9 10 11
12 13 14 15 16 17 18
19 20 21 22 23 24 25
26 27 28 29 30 31
 

30 July 2009

Tim Weenink

prepared assembly mixes for !B, !C and !D constructs. Standard Ginko assembly kit was used. DNA concentrations as below (these mixtures were added to 7.5 µl of digestion mixture):

Assembly name position Components volume in µl
 !B Upstream  !A DNA 33.0
H20 9.5
Downstream BBa_B0015 DNA 10.0
H20 32.5
Backbone pSB1AC3 DNA 6.5
H20 36
 !C Upstream BBa_R0040 DNA 10.0
H20 32.5
Downstream *I6 DNA 5.5
H20 37.0
Backbone pSB1AC3 DNA 6.5
H20 36
 !D Upstream *I6 DNA 5.5
H20 37.0
Downstream BBa_K145201 DNA 18.0
H20 24.5
Backbone pSB1AC3 DNA 6.5
H20 36

Ligated and transformed assemblies according to protocol.

15:20 Done plating the assemblies.

15:40 inoculated *S and *T cultures, four colonies each

16:55 Inoculated !A cultures from glycerole stock. Two tubes.

Calin

Plated three backbones left over from the transformation the other day. Did a digest on oriT-R with E and P. Made 3 x 5ml culture tubes for the pSB4C5 backbone which had a few colonies. Made electrocompetent cells with the Knight:Preparing electrocompetent cells protocol. Flash froze tube in liquid nitrogen. Checked backbone concentrations:


Part Concentration (ng/uL)
rbs-GFP-term I 102.6
rbs-GFP-term II 113.5
pSB1A3 I 89.1
pSB1A3 II 83.2
pSB1A3 III 91.9
pSB1A3 IV 84.7
pSB4A5 I 77.7
pSB4A5 II 83.9
pSB4A5 III 99.0
pSB4A5 IV 116.1
pSB1C3 I 102.3
pSB1C3 II 49.4
pSB1C3 III 81.5
pSB1C3 IV 66.0

Ran gel on backbones and oriT digest. Gel left for too long.

Gel30072009.png


Well Part Plasmid Size
1 rbs-GFP-term I
2 rbs-GFP-term II
3 pSB1A3 I 2832
4 pSB1A3 II 2832
5 pSB1A3 III 2832
6 pSB1A3 IV 2832
7 pSB4A5 I 4070
8 pSB4A5 II 4070
9 pSB4A5 III 4070
10 pSB4A5 IV 4070
11 DNA Ladder
12 pSB1C3 I 2747
13 pSB1C3 II 2747
14 pSB1C3 III 2747
15 pSB1C3 IV 2747
16 oriT E + P digest
17 oriT E + P digest
18 DNA Ladder

Tim Vos

Minipreped 4 x pSB1C3, 4 x pSB4A5, 2 x rbs-GFP, and 4 x pSB1A3. One pSB4A5 tube is incorrect, it shows RFP expression. Also loaded the gel.

Orr

Made some chemically competent cells using the TSS buffer protocol. Started making a Java program that will allow lock sequence design based on the chosen RBS, and will design the equivalent key based on the lock sequence. The program will incorporate a database with all the working RBS segments in the iGEM registry, so that the user can choose which RBS he wants to use.


Retrieved from "http://2009.igem.org/Team:TUDelft/30_July_2009"