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Cell lysis buffer for real time RT-PCR - Request for cell lysis buffer recipe or relevant scientific paper (Feb/20/2008 )

Hey

Does any know of a cell lysis buffer that is compatible with downstream real time RT-PCR analysis? That is, a buffer that lyse the cells and which can be used directly for reverse transcription so no RNA isolation is required. If anyone would be willing to share such a buffer recipe or has come across a relevant paper/reference I would very much appreciate it.

I am aware that such a lysis buffer is commercially available from, forexample, Ambion (Cells-to-cDNA™ II Cell Lysis Buffer) but it is somewhat expensive.

-Potok-

QUOTE (Potok @ Feb 20 2008, 08:50 AM)
Hey

Does any know of a cell lysis buffer that is compatible with downstream real time RT-PCR analysis? That is, a buffer that lyse the cells and which can be used directly for reverse transcription so no RNA isolation is required. If anyone would be willing to share such a buffer recipe or has come across a relevant paper/reference I would very much appreciate it.

I am aware that such a lysis buffer is commercially available from, forexample, Ambion (Cells-to-cDNA™ II Cell Lysis Buffer) but it is somewhat expensive.


Make a solution of 10 mM glycine, 1% Triton X-100, pH to 2.5 with HCl. Wash your cells with room temp. PBS, then add this buffer, also at room temp. Allow it to sit on the cells for ~2 minutes, then pipet up-and-down until the cells come off (this will vary by cell line). For a 96-well plate, use 100 uL of the buffer, and scale accordingly.

-alydar-