Team:TUDelft/6 August 2009

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='''6 August 2009'''=
='''6 August 2009'''=
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===Sriram===
 +
Today I continued with the ligation of the restricted parts done yesterday. Then the ligated parts were electro transformed and also heat shock transformed. There are in total 8 assembly as follows: <br>
 +
1. pTet + RBS-cI-RBS (Chloramphenicol backbone) <br>
 +
2. mRFP + Double Terminator (Chloramphenicol backbone) <br>
 +
3. pLacI + RBS (Chloramphenicol backbone) <br>
 +
4. TetR + Double Terminator (Chloramphenicol backbone) <br>
 +
5. pTet + Lock3c (Chloramphenicol backbone) <br>
 +
6. cI + Double Terminator (Chloramphenicol backbone) <br>
 +
7. &lambda;p-in + RBS-GFP-Double Term (Chloramphenicol backbone) <br>
 +
8. pLacI + key3c (Amp-Kan backbone) <br>
 +
 +
===Calin===
 +
 +
Good results on the conjugation test. Many colonies. All plates were imaged on the safe imager with the 5 Megapixel camera. The plates with GFP (T and R) were excited by blue light and placed under a filter (50 ms exposure). The plates without GFP (D) were imaged with the upper white light and an exposure of 15ms. Images were color inverted before the image analysis. Colony counting was done with [http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1770926 Clono-Counter].
 +
 +
 +
 +
{| border="1" align="center"
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|-
 +
| <b>Set 1</b>
 +
|- align="center"
 +
| Plate ID || Antibiotics || Dilution || # Colonies
 +
|- align="center"
 +
| D1 || TRI || 10<sup>0</sup> || overgrown
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-D2-noGFP.jpg D2] || TRI || 10<sup>-2</sup>|| <font color=green><B>6770</b></font>
 +
|- align="center"
 +
| D3 || TRI || 10<sup>-4</sup>|| 170
 +
|- align="center"
 +
| D4 || TRI || 10<sup>-5</sup>|| 40
 +
|- align="center"
 +
| D5 || TRI || 10<sup>-6</sup>|| 10
 +
|- align="center"
 +
| T1 || TRI + AMP || 10<sup>0</sup> || ?
 +
|- align="center"
 +
| T2 || TRI + AMP || 10<sup>-1</sup>|| ?
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-T3-GFP.jpg T3] || TRI + AMP || 10<sup>-2</sup>|| <font color=green><B>186</b></font>
 +
|- align="center"
 +
| T4 || TRI + AMP || 10<sup>-3</sup>|| 11
 +
|- align="center"
 +
| T5 || TRI + AMP || 10<sup>-4</sup>|| 0
 +
|- align="center"
 +
| T6 || TRI + AMP || 10<sup>-5</sup> || 0
 +
|- align="center"
 +
| T7 || TRI + AMP || 10<sup>-6</sup>|| 0
 +
|- align="center"
 +
| R1 || AMP || 10<sup>0</sup> || overgrown
 +
|- align="center"
 +
| R2 || AMP || 10<sup>-4</sup>|| 32
 +
|- align="center"
 +
| R3 || AMP || 10<sup>-5</sup>|| 6
 +
|- align="center"
 +
| R4 || AMP || 10<sup>-6</sup>|| 2
 +
|}
 +
 +
<br><br>
 +
{| border="1" align="center"
 +
|-
 +
| <b>Set 2</b>
 +
|- align="center"
 +
| Plate ID || Antibiotics || Dilution ||
 +
|- align="center"
 +
| D6 || TRI || 10<sup>0</sup> || overgrown
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-D7-noGFP.jpg D7] || TRI || 10<sup>-2</sup>|| <font color=green><B>8320</b></font>
 +
|- align="center"
 +
| D8 || TRI || 10<sup>-4</sup>|| 114
 +
|- align="center"
 +
| D9 || TRI || 10<sup>-5</sup>|| 18
 +
|- align="center"
 +
| D10 || TRI || 10<sup>-6</sup>|| 8
 +
|- align="center"
 +
| T8 || TRI + AMP || 10<sup>0</sup> || ?
 +
|- align="center"
 +
| T9 || TRI + AMP || 10<sup>-1</sup>|| ?
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-T10-GFP.jpg T10] || TRI + AMP || 10<sup>-2</sup>|| <font color=green><B>228</b></font>
 +
|- align="center"
 +
| T11 || TRI + AMP || 10<sup>-3</sup>|| 18
 +
|- align="center"
 +
| T12 || TRI + AMP || 10<sup>-4</sup>|| 2
 +
|- align="center"
 +
| T13 || TRI + AMP || 10<sup>-5</sup> || 0
 +
|- align="center"
 +
| T14 || TRI + AMP || 10<sup>-6</sup>|| 0
 +
|}
 +
 +
<br><br>
 +
{| border="1" align="center"
 +
|-
 +
| <b>Set 3</b>
 +
|- align="center"
 +
| Plate ID || Antibiotics || Dilution ||
 +
|- align="center"
 +
| D11 || TRI || 10<sup>0</sup> || overgrown
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-D12-noGFP.jpg D12] || TRI || 10<sup>-2</sup>|| <font color=green><B>8950</b></font>
 +
|- align="center"
 +
| D13 || TRI || 10<sup>-4</sup>|| 173
 +
|- align="center"
 +
| D14 || TRI || 10<sup>-5</sup>|| 22
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|- align="center"
 +
| D15 || TRI || 10<sup>-6</sup>|| 1
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|- align="center"
 +
| T15 || TRI + AMP || 10<sup>0</sup> || ?
 +
|- align="center"
 +
| T16 || TRI + AMP || 10<sup>-1</sup>|| ?
 +
|- align="center"
 +
| [https://2009.igem.org/Image:TUDelft-aug5-2009-conj-test-plate-T17-GFP.jpg T17] || TRI + AMP || 10<sup>-2</sup>|| <font color=green><B>218</b></font>
 +
|- align="center"
 +
| T18 || TRI + AMP || 10<sup>-3</sup>|| 31
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|- align="center"
 +
| T19 || TRI + AMP || 10<sup>-4</sup>|| 2
 +
|- align="center"
 +
| T20 || TRI + AMP || 10<sup>-5</sup> || 0
 +
|- align="center"
 +
| T21 || TRI + AMP || 10<sup>-6</sup>|| 0
 +
|}
 +
 +
<br><br>
 +
Calculated conjugation efficiencies:
 +
 +
{| border="1" align="center"
 +
|- align="center"
 +
| Set || Conjugation Efficiency
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|- align="center"
 +
| 1 ||  0.028
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|- align="center"
 +
| 2 ||  0.028
 +
|- align="center"
 +
| 3 ||  0.025
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|}
 +
 +
Trimethoprim control plate had no growth.
 +
 +
5mL tube of assembly CA made.
 +
 +
Elecroporated 10 assemblies. Experienced some arcing with assemblies 1 and 6. Assembly 6 was redone with only 2 uL of DNA added.
 +
 +
===Tim Weenink===
 +
 +
Elecroporated 10 assemblies.
 +
 +
{| border="1" align="center"
 +
|- align="center"
 +
| Biobrick || origin of parts || 
 +
|}
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 +
===Orr===
 +
Made 2 one liter batches of LB medium, and prepared some chemically competent TSS cells.
 +
 +
[[Image:Tim060809!D.png|200px|]]
{{Template:TUDelftiGEM2009_end}}
{{Template:TUDelftiGEM2009_end}}

Latest revision as of 08:01, 20 October 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
 

6 August 2009

Sriram

Today I continued with the ligation of the restricted parts done yesterday. Then the ligated parts were electro transformed and also heat shock transformed. There are in total 8 assembly as follows:
1. pTet + RBS-cI-RBS (Chloramphenicol backbone)
2. mRFP + Double Terminator (Chloramphenicol backbone)
3. pLacI + RBS (Chloramphenicol backbone)
4. TetR + Double Terminator (Chloramphenicol backbone)
5. pTet + Lock3c (Chloramphenicol backbone)
6. cI + Double Terminator (Chloramphenicol backbone)
7. λp-in + RBS-GFP-Double Term (Chloramphenicol backbone)
8. pLacI + key3c (Amp-Kan backbone)

Calin

Good results on the conjugation test. Many colonies. All plates were imaged on the safe imager with the 5 Megapixel camera. The plates with GFP (T and R) were excited by blue light and placed under a filter (50 ms exposure). The plates without GFP (D) were imaged with the upper white light and an exposure of 15ms. Images were color inverted before the image analysis. Colony counting was done with Clono-Counter.


Set 1
Plate ID Antibiotics Dilution # Colonies
D1 TRI 100 overgrown
D2 TRI 10-2 6770
D3 TRI 10-4 170
D4 TRI 10-5 40
D5 TRI 10-6 10
T1 TRI + AMP 100  ?
T2 TRI + AMP 10-1 ?
T3 TRI + AMP 10-2 186
T4 TRI + AMP 10-3 11
T5 TRI + AMP 10-4 0
T6 TRI + AMP 10-5 0
T7 TRI + AMP 10-6 0
R1 AMP 100 overgrown
R2 AMP 10-4 32
R3 AMP 10-5 6
R4 AMP 10-6 2



Set 2
Plate ID Antibiotics Dilution
D6 TRI 100 overgrown
D7 TRI 10-2 8320
D8 TRI 10-4 114
D9 TRI 10-5 18
D10 TRI 10-6 8
T8 TRI + AMP 100  ?
T9 TRI + AMP 10-1 ?
T10 TRI + AMP 10-2 228
T11 TRI + AMP 10-3 18
T12 TRI + AMP 10-4 2
T13 TRI + AMP 10-5 0
T14 TRI + AMP 10-6 0



Set 3
Plate ID Antibiotics Dilution
D11 TRI 100 overgrown
D12 TRI 10-2 8950
D13 TRI 10-4 173
D14 TRI 10-5 22
D15 TRI 10-6 1
T15 TRI + AMP 100  ?
T16 TRI + AMP 10-1 ?
T17 TRI + AMP 10-2 218
T18 TRI + AMP 10-3 31
T19 TRI + AMP 10-4 2
T20 TRI + AMP 10-5 0
T21 TRI + AMP 10-6 0



Calculated conjugation efficiencies:

Set Conjugation Efficiency
1 0.028
2 0.028
3 0.025

Trimethoprim control plate had no growth.

5mL tube of assembly CA made.

Elecroporated 10 assemblies. Experienced some arcing with assemblies 1 and 6. Assembly 6 was redone with only 2 uL of DNA added.

Tim Weenink

Elecroporated 10 assemblies.

Biobrick origin of parts

Orr

Made 2 one liter batches of LB medium, and prepared some chemically competent TSS cells.

Tim060809!D.png

Retrieved from "http://2009.igem.org/Team:TUDelft/6_August_2009"