inhibitors of RNA & protein synthesis.

Kevin Seeley kseeley at NATURE.BERKELEY.EDU
Thu Jan 4 11:55:53 EST 1996

On 3 Jan 1996, Chris Town wrote:

> I would like to inhibit new gene expression in some experiments designed to 
> investigate the inducibility of a hormone conjugation system. 
> I would appreciate advice of the effectivenss of traditional eukaryotic 
> inhibitors such as actinomycin D, alpha-amanitin, cycloheximide.
> Test material will be seedlings in suspension.
> Do these compounds work? What concentrations? How effective?
> References?
> I'll summarize and post any relies that don't appear directly on the net.
> Thank y'all for your time.
> Happy New Year.
> Chris Town

To Chris Town 

See:    Seeley et al., 1991, 
        Red light-independant instability of oat phytochrome mRNA in vivo
        Plant Cell 4:29-38

Also:   Bryne et al., 1993,
        Half-lives of oat mRNA's in vivo and in a polysomal-based in-vitro 
        system.  Planta  189:249-256.

I used cordycepin to prevent new synthesis of mRNA in order to quantify the
mRNA half-life of phytochrome mRNA in oat seedlings.  The seedlings were
suspended in a buffer solution containing the inhibitors at various
concentrations for periods of 1 to 4 hrs following pretreatment. We used a
light inducible message as a control to verify that mRNA systhesis was
fully inhibited.  A paper by Edelmann and Schopfer served as a guide for
our initial setup, they were performing auxin treatments on inhibitor
treated oat coleptiles (referenced in the Seeley et al., paper).  In
unpublished work, I used cycloheximide (superinduction of seady state mRNA
levels at 100ug/ml) and alpha-amanitin (did not seem to inhibit well even
at high concentrations).

Good Luck

Dr. Kevin Seeley                                        TEL     510-642-0404
111 Koshland Hall
Plant Biology/SUNG                                      FAX     510-642-4995
University of California
Berkeley, CA  94720

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