Uppsala-Sweden/31 July 2009

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OD values were measured in triplicates.  
OD values were measured in triplicates.  
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Results here:[[Media:1st_read_2009_07_31.xls]]
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Results here: [[Media:1st_read_2009_07_31.xls]]
--[[User:Karl.brune|Karl.brune]] 16:38, 31 July 2009 (UTC)
--[[User:Karl.brune|Karl.brune]] 16:38, 31 July 2009 (UTC)
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Thanks to Fernando for helping us, handling the slimy fil.
Thanks to Fernando for helping us, handling the slimy fil.
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As mentioned by Karl we managed to extract DNA from the Verum Hälsofil and Långfil using the protocol ''Protocol Ethanol-Filmjölkl.doc'' at [[http://uppsalaigem.pbworks.com/browse/#view=ViewFolder&param=Protocols]] (unavailable for non users). In the last separation step we didn't get the pellet as desired and thus moved on to the alternative route mentioned in the protocol. In the end we managed to extract DNA from both of the products which we then stored in the freezer awaiting PCR.
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--[[User:Anders|Anders]] 19:45, 1 August 2009 (UTC)
--[[User:Karl.brune|Karl.brune]] 16:38, 31 July 2009 (UTC)
--[[User:Karl.brune|Karl.brune]] 16:38, 31 July 2009 (UTC)
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== Preparing competent E.coli ==
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''continued since 30th of July''
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Today I finished the preparation of competent cells, in total 120 eppendorf tubes with 100 µl each was produced. This was done according to the protocol [http://openwetware.org/wiki/TOP10_chemically_competent_cells ] with the following exceptions:
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The OD in the growing step reached 0,460. The cells grew faster then anticipated. Diluted from 300 ml to 500 ml waited and then moved on.
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To in part compensate the larger volume I harvested 6 x 50 ml (300 total) of cell suspension in 50 ml falcon tubes which was used in the centrifugation instead of flat bottomed centrifugation bottles. This necessitated more CCMB80 washing buffer in the first washing step, after which I transferred all suspension to two tubes.
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No OD measurement of the OD was performed after washing as this was deemed unnecessary - we'll test the competence in any case.
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Finally the cells were transferred to eppendorf tubes, 100 µl each, and stored at - 80 degrees C.
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--[[User:Anders|Anders]] 20:00, 1 August 2009 (UTC)

Latest revision as of 20:00, 1 August 2009




Does Z. Mobilis kill Synechocystis?

As our prelimary test from the ethanol evolution appear to show that Z. mobilis and Synechocystis do not like to be grown together, we decided to make a small side project, to test if Z. mobilis really kills Synechocystis. Therefore we repeated the expirement done in the ethanol evolution project with triplicates.

Triplicates of 14,5 ml BG11 + 0,69 ml EtOH (thus 4%) + 1ml Synchecocystis inoculum + 0,5ml Z.mobilis inoculum are incubated at 30°C in the cyanobacteria growth room. Control is 14,5 ml BG11 + 0,69 ml EtOH (thus 4%) + 1ml Synchecocystis inoculum + 0,5ml BG11.

Triplicates of 14,5 ml BG11 + 1ml Synchecocystis inoculum + 0,5ml Z.mobilis inoculum are incubated at 30°C in the cyanobacteria growth room. Control is 14,5 ml BG11 + 1ml Synchecocystis inoculum + 0,5ml BG11.

OD values were measured in triplicates. Results here: Media:1st_read_2009_07_31.xls

--Karl.brune 16:38, 31 July 2009 (UTC)

DNA-Extraction from Fil products

DNA-Extraction: Anders and I made further DNA extraction from Hälsofil and Langfil to use them as templates for our PCR for the kivd genes. Protocol is coming soon.

Thanks to Fernando for helping us, handling the slimy fil.


As mentioned by Karl we managed to extract DNA from the Verum Hälsofil and Långfil using the protocol Protocol Ethanol-Filmjölkl.doc at http://uppsalaigem.pbworks.com/browse/#view=ViewFolder&param=Protocols (unavailable for non users). In the last separation step we didn't get the pellet as desired and thus moved on to the alternative route mentioned in the protocol. In the end we managed to extract DNA from both of the products which we then stored in the freezer awaiting PCR.

--Anders 19:45, 1 August 2009 (UTC)

--Karl.brune 16:38, 31 July 2009 (UTC)

Preparing competent E.coli

continued since 30th of July

Today I finished the preparation of competent cells, in total 120 eppendorf tubes with 100 µl each was produced. This was done according to the protocol [http://openwetware.org/wiki/TOP10_chemically_competent_cells ] with the following exceptions:

The OD in the growing step reached 0,460. The cells grew faster then anticipated. Diluted from 300 ml to 500 ml waited and then moved on.

To in part compensate the larger volume I harvested 6 x 50 ml (300 total) of cell suspension in 50 ml falcon tubes which was used in the centrifugation instead of flat bottomed centrifugation bottles. This necessitated more CCMB80 washing buffer in the first washing step, after which I transferred all suspension to two tubes.

No OD measurement of the OD was performed after washing as this was deemed unnecessary - we'll test the competence in any case.

Finally the cells were transferred to eppendorf tubes, 100 µl each, and stored at - 80 degrees C.

--Anders 20:00, 1 August 2009 (UTC)

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