Team:UNICAMP-Brazil/Protocols/Preparation of electrocompetent S. cereviseae
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(New page: ==Prepare of eletrocompeten ''S. serevisiae''== 1- Grow yest strain in 50ml YEPD medium overnight, 250 RPM, 30ºC. 2- Inoculate 100 ml YEPD in a 500ml flask in a OD=0,1, 250 RPM, 30...)
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(New page: ==Prepare of eletrocompeten ''S. serevisiae''== 1- Grow yest strain in 50ml YEPD medium overnight, 250 RPM, 30ºC. 2- Inoculate 100 ml YEPD in a 500ml flask in a OD=0,1, 250 RPM, 30...)
Newer edit →
Revision as of 14:19, 20 October 2009
Prepare of eletrocompeten S. serevisiae
1- Grow yest strain in 50ml YEPD medium overnight, 250 RPM, 30ºC.
2- Inoculate 100 ml YEPD in a 500ml flask in a OD=0,1, 250 RPM, 30ºC util OD= 1,3
3- Place the culture on ice for 15 minutes to stop the growth
4- Centrifuge for 4 min at 4000g, 4ºC
5- Ressuspend cells in 20ml of cold water in 50ml tubes, complete volum to 50ml.
6- Centrifuge for 4 min at 4000g, 4ºC
7- Repeat steps 5 and 6.
8- Ressuspend cells in 10ml Sorbitol 1M
9- Centrifuge for 4 min at 4000g, 4ºC
10- Ressuspend cells in 200ul
DNA extraction