Real-Time PCR using genomic DNA (without DNA purification)
Posted 29 April 2010 - 10:55 AM
• Add to the cell pellets:
- 100 µl Kawasaki buffer (20 mM Tris HCl, 1.5 mM MgCl2 , 25 mM KCl, 0.5% Tween-20 (v/v))
- 6 µl proteinase K (20 mg ml-1, dissolved in aqua bidest)
• Incubate the samples at 56 °C overnight (cell lysis).
• heat the samples to 95°C for 15 min to inactivate proteinase K.
• centrifuge at 15300g and 4° C for 5 min
• Use the lysed samples directly in the PCR analysis without any further manipulations.
(Harald Lahm et al.. Transgenic Research 7, 131-134 (1998))
Do you know wich kit for the analysis of GAPDH expression can I use for the normalization of my samples?
I found some kits from QIAGEN (SeqTarget LongRange PCR), but I am sure that someone knows a better solution for me.
Posted 30 April 2010 - 12:27 AM
Posted 25 May 2011 - 04:48 PM