Team meeting

1. presentations of work did by both groups during last week (given by Ania and Kuba)
2. presentation of methods used by teams starting in previous editions of iGEM to kill bacteria :) (Kamil)
3. presentation of different methods of targeting drugs to cancer cells (Marcin)

We've been also talking about some organization issues and we've decided to move our meetings - now they will take place every Thursady at 17:00, one week at the Faculty of Biology Building, another week in the room E of the lab on Pawinskiego Street.

Creating devices to test promoters in E. coli strains (devices [http://partsregistry.org/Part:BBa_K177024 BBa_K177024] and [http://partsregistry.org/Part:BBa_K177025 BBa_K177025])

Franek

Tasks:

• Alkaline lysis of the plasmid containing [http://partsregistry.org/Part:BBa_B0024 BBa_B0024 terminator]

Methods:

• [http://aabiot.com/products/dna_purification/plasmid_dna/plasmid_mini/protocol_plasmid_mini.pdf PlasmidMini set by A&A Biotechnology] was used. 2 test tubes with 5ml LB and ampicillin were first inoculated with colonies containing [http://partsregistry.org/Part:BBa_B0024 BBa_B0024] brick on [http://partsregistry.org/Part:pSB1A2 pSB1A2] plasmid. The cultures were incubated for 6h at 37°C. Alkaline lysis was performed on both cultures. The pellet from 5 ml of bacteria was used.

Results:

• Concentration of DNA sample was measured using NanoDrop ND-1000

Notes:

• Plates with [http://partsregistry.org/Part:BBa_I0500 BBa_I0500] transformants were empty once again. This result, together with note in [http://partsregistry.org/partsdb/get_part.cgi?part=BBa_I0500 part description] suggest that this part is damaged

Making of RBS-cI part

Jarek

Task:

• Digestion of parts C0051 and B0032
• Electrophoretic separation of digested parts
• Isolation of DNA samples from gel
• Ligation of part C0051 to the vector with B0032

Methods:

• DNA samples were digesteg with 1 µl of PstI/BcuI (B0032) or PstI/XbaI (C0051) in 1xTango buffer for 3 hours.
• After digestion they were separated on 0,8% agarose gel.
• Isolation of sample from gel was performed with the A&A "Gel-out" kit.
• For ligation 4µl of ligase buffer and 2 µl of ligase were used.

Results:

Cloning of p53 coding sequence

Marcin

Comment:

Due to force problem with low quality of PCR product it was necessary to transform bacteria with plasmid containing p53 sequence and to perform all procedures once more time

Tasks:

• Transformation of chemocompetent strain of E. coli by plasmid containing p53 sequence

Procedure:

• The protocol of transformation has not been changed, except of giving 1 µl of plasmid solution to the bacteria. If you want to see detailed procedure go here
• Bacteria was plated on the medium containing kanamycin

Construction of [http://partsregistry.org/wiki/index.php?title=Part:BBa_K177012 K177012] operon1_part2

Ania

Tasks:

• Competent cells were transformed with LacI C0012 taken out of the distribution 2009 Kit Plate 1 well 20.

Isolation of BioBricks from 2008 and 2009 Kit Plates

Monika

Tasks:

• Isolate plasmids containing the biobricks - continuation

1. GFP coding device switched on by IPTG - [http://partsregistry.org/Part:BBa_I763004 BBa_I763004] from 2008 Kit Plate 1017 well G5
2. promoter lambda (cI regulated) with RFP reporter [http://partsregistry.org/Part:BBa_I763007 BBa_I763007] from 2009 Kit Plate 1 well 15J
3. GFP generator - [http://partsregistry.org/Part:BBa_E0840 BBa_E0840] from 2009 Kit Plate 1 well 12O
4. GFP generator - [http://partsregistry.org/Part:BBa_J07037 BBa_J07037] from 2009 Kit Plate 1 well 12O

Results from 9 July 2009

• all transformations exept GFP coding device switched on by IPTG (BBa_I763004) were successful

Methods:

• Planting on LB medium supplemented with apropriate antibiotic, 6h incubation in 37°C
• Planting on LB-agar medium supplemented with apropriate antibiotic
• Alkaline lysis (prodedure described [http://www.aabiot.com/products/dna_purification/plasmid_dna/plasmid_mini/protocol_plasmid_mini.pdf here])

Results

• Concentration of DNA sample was measured using NanoDrop ND-1000

Comment

• Isolations where DNA concentration is under 30ng/µl must be repeated