RNA isolation from PANCREAS!!!
bbecker at iastate.edu
Thu Dec 1 11:41:52 EST 1994
In article <D0371C.8wI at liverpool.ac.uk>,
Mr J.S. Erikson <ferret at liverpool.ac.uk> wrote:
> Am I having major problems with this one.
>Ive been isolating RNA from tissues and from leukocytes using several diff
>erent methods; Trizol(Gtc complex [GIBCO], Lithium chloride/urea; and alkaline
>lysis methods. Recently Ive tried to isolate total RNA, with the hope of
>performing Northern hybridisation from PANCREAS.
>Q. The surgeon who is excising the pancreas has been snap freezing them
>in liquid nitrogen, will this fragment mRNAs of approx 1-2kb?
>Q. Would it help to excise the pancreas and immediatley homogenise in
>the isolation agent?
> Unfortunately Im losing the RNA through degredation. Ive had an internal
>control in each extraction (Human P.B.L.s) to check my technique is ok.
> Any good ideas out there? e-mail: ferret at liv.ac.uk
> Thanks people John Erikson
Dear Mr. Erikson,
The isolation of intact RNA from pancreatic tissue is probably
one of the most tricky RNA isolations due to the high RNAse content of
pancreas. However, one method has been published that yields high
quality RNA even from pancreas. I have been taught this method by one of
the authors of the paper (Dr. C. Stratowa), and it has always worked
fine for me.
The paper is:
Isolation of full-length putative rat lysophospholipase cDNA using
improved methods for mRNA isolation and cDNA cloning. J.H. Han, C.
Stratowa, and W.J. Rutter. Biochemistry 1987, 26, pp. 1617-1625.
In case you would need to know some details of this procedure,
please let me know.
Email: bbecker at iastate.edu
Phone (USA): (515)294-3658
Fax (USA): (515)294-4141
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