Plasmid DNA purification
Posted 03 September 2004 - 04:36 AM
We are looking for a plasmid purification technique (kit or manual) that fulfills the following criteria:
1) begins with an approximate 50 mL culture
2) can prep 20 at a time, with or without vacuum, or spin
3) give DNA clean enough for electroporation
4) doesn't use phenol
We find that a typical alkaline lysis procedure from 1.5 mL culture of Miniprep DNA give about 40 ug of DNA; yet Qiagens plasmid preps (and Sigma which we have also tried) have given us less than 100 ug cumulatively when we do 50 mL preps. This means that column purifications are very inefficient. Can one scale up Maniatis efficiently, for example?
Posted 03 September 2004 - 11:18 AM
Posted 05 September 2004 - 09:07 PM
The quality and yield (600-700 ug per 200-300 mL culture) are astonishing and very satisfying. Most importantly, the price is affordable and much less than Qiagen's. If you're interested, check their web site www.epochbiolabs.com
Posted 14 September 2004 - 07:57 AM
It is working just fine, it fullfills the criteria, but i do not know
how many samples you can run at the same time.
Maybe not 20...
Posted 15 September 2004 - 03:00 AM
Posted 05 January 2005 - 09:24 AM
Posted 05 January 2005 - 09:48 AM
I wonder how u get 40ug of Plasmid DNA from 1.5 ml culture. I never got that much.
If u allow me I would like to use your protocol.
Posted 06 January 2005 - 02:42 PM
I work for Promega's Technical Services deprtment. The requiremtns you have meet the specifications of our new PureYield™ Plasmid Midiprep system. Witha vacuum manifold exactly 20 samples can be processed at one time from 50mL of culture and we typically see well over 200ug of DNA. Using the link beow will bring you to a survey that allows you to receive a free trial of the system.
If you have any additional questions, please feel free to contact me here, or preferably by e-mail at firstname.lastname@example.org