RNA degradation in collagen gels (View forum version)


Posted 09 November 2010 - 03:49 AM

Hi there, i have been trying to extract total RNA from collagen gels and no matter which way i disrupt the membrane i get degraded RNA, i have tried liquid N2 disruption, gentleMACS dissociation and simply cutting it up and putting it in TRIZOL. my cell line positive control done at the same time, equipment etc is absolutely fine with ribosomal bands on agarose gel. Was wandering if anyone had any similar experience or suggestions?


Posted 09 November 2010 - 09:48 AM

My first suggestion is going to be to make sure you're using proper RNA handling technique - Try to do everything at 4*C, make sure everything that will be touching your sample is RNase-free or cleaned with RNase-away / RNaseZap before use, wear gloves, try not to move your hands or clothing above open samples, don't breathe on them, etc...

And also, if I'm reading you correctly, you're assaying for total RNA in your sample but looking for rRNA in your control? Why would you treat the two differently? You should assay for the same type of RNA in both.