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Uppsala-Sweden/4 August 2009
From 2009.igem.org
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==Butanol Evolution== | ==Butanol Evolution== | ||
26ml flasks from the Butanol recovery were supplied with isobutanol regarding their former concentration. (0,25% | 0,25% | 0,5% | control w/o). Flask were incubated in 30°C growth room on 120rpm shaker. | 26ml flasks from the Butanol recovery were supplied with isobutanol regarding their former concentration. (0,25% | 0,25% | 0,5% | control w/o). Flask were incubated in 30°C growth room on 120rpm shaker. | ||
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| + | ==New Pir A Primers== | ||
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| + | New Primers for the Pir A gene was ordered | ||
| + | [[Media:PirA primers v_2.doc]] | ||
==PCR (Kivd, ADH2 and PpetE) and Gel run== | ==PCR (Kivd, ADH2 and PpetE) and Gel run== | ||
Revision as of 12:03, 5 August 2009
Contents |
Ethanol Evolution
1,15ml of 95 EtOH were added to a 26ml flask from the recovery protocol of the ethanol evolution project. Thus resulting in approx. 4% EtOH concentration (0,03981% in theory) but I expect that some water evaporated during the culturing thus the former volume (26ml) was decreased. Flask were incubated in 30°C growth room on 120rpm shaker.
Butanol Evolution
26ml flasks from the Butanol recovery were supplied with isobutanol regarding their former concentration. (0,25% | 0,25% | 0,5% | control w/o). Flask were incubated in 30°C growth room on 120rpm shaker.
New Pir A Primers
New Primers for the Pir A gene was ordered Media:PirA primers v_2.doc
PCR (Kivd, ADH2 and PpetE) and Gel run
This is continued since yesterday.
We ran the PCR products on a gel with the following setup and result.
Setup: Media:gel_run_090804_adh2_kivd_ppete.xls
Result:
As clearly seen the PCR failed for all but the PpetE. For kivd the straightforward explanation is absence of the correct strain of lactococcus, although reasons such as poor preparation cannot be ruled out due to the lack of a control (bad planning). In the case of ADH2 a likely cause of failure is the toughness of the yeast cell, preventing exposure of the genome without special lysis being performed.
Thus we will try to find other sources for the correct Lactococcus and repeat the adh2 after lysis of the yeast cells. --Flormane 12:32, 4 August 2009 (UTC)
PCR for transformation
genes: PDC,ADH2 from z.mobilis, PirB, ADH2 from Bakers/Wine yeast and PpetE)
A PCR run was started to make products for the transformation.
This was the setup of annealing Tm and primer design: Media:primers_090804_adh2z_adh2y_pirb_pdc_ppete.xls --Flormane 16:47, 4 August 2009 (UTC)
