Neurospora RNA for RNase protection

Matthew Sachs msachs at ADMIN.OGI.EDU
Mon Mar 13 15:45:29 EST 1995


>Is anyone using a particular RNA isolation protocol for Neurospora which
>yields total RNA suitable for RNase protection?  I seem to be having trouble
>with DNA contamination using techniques published in the FGN.
>
>Ideally, I would like a technique which is quick, easy and doesn't involve
>isolating mRNA from total.  Is this wishful thinking?
>
>If you have any suggestions, please email me.  Thanks in advance.
>
>Martin Hoyt
>Dept. of Molecular Biology and Biochemistry, Univ. Calif. Irvine
>mahoyt at uci.edu


1.  Lower pH (5-6) in aqueous buffers for RNA extraction is reported to
reduce the recovery of contaminating DNA.

2.  A simple but not quantitative method to separate DNA (and small RNA)
from larger RNA, such as mRNA, is on the basis of their differential
solubilities in lithium chloride.

3. RNase-free DNase can be used to treat total RNA samples.

See:

Wallace, D. M. (1987). Large- and small-scale phenol extractions. Meth.
Enzymol. 152, 33-41.

Wallace, D. M. (1987). Precipitation of nucleic acids. Meth. Enzymol. 152,
41-48.



-----------------------------------------------------------
Matthew Sachs
Department of Chemistry, Biochemistry and Molecular Biology
Oregon Graduate Institute of Science and Technology
20000 NW Walker Road
P.O. Box 91000
Portland, OR  97291-1000
503 690-1487 Phone
503 690-1464 Fax
msachs at admin.ogi.edu





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