Waldminghaus:Back Door/Sonication: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(New page: =Sonication= Decide wether you would like to sonicate in a 15 ml Falcon tubes or in a 1,5 ml Eppendorf tube. In a 15 ml Falcon tube you can only fill in a maximal volume of 1,7 ml. In ...) |
mNo edit summary |
||
| Line 1: | Line 1: | ||
=Sonication= | =Sonication= | ||
Decide wether you would like to sonicate in a 15 ml Falcon | Decide wether you would like to sonicate in a 15 ml Falcon tube or in a 1,5 ml Eppendorf tube. | ||
In a 15 ml Falcon tube you can only fill in a maximal volume of 1,7 ml. | In a 15 ml Falcon tube you can only fill in a maximal volume of 1,7 ml. | ||
Latest revision as of 11:09, 3 November 2012
Sonication
Decide wether you would like to sonicate in a 15 ml Falcon tube or in a 1,5 ml Eppendorf tube.
In a 15 ml Falcon tube you can only fill in a maximal volume of 1,7 ml. In an 1,5 ml Eppendorf tube you can only fill in a maximal volume of 300 ul (otherwise the sonication will not work out very well).
Procedure
- Resuspend your bacteria pellet in IP buffer.
- Sonicate for 30 s at high level i.e. 48 times in Eppendorf tubes if you wish to have chromosomal E. coli DNA at a size of around 200 – 500 bp.
- Cool the samples for 30 s between the 48 cycles.
- After sonication centrifuge for 10 minutes, 4°C at around 7150 g.
- Take the supernatant (= sheared DNA) and then go on with your further procedure.
