lab protocol for GA induction of amylase

Dr. David Starrett dstarret at BIOLOGY.SEMO.EDU
Fri Feb 13 11:03:25 EST 1998


>
>        Can anyone help out? I've got a pretty good idea of what to expect,
>but need to know such things as - how long does it take to go from dry
>seeds to a enough starch breakdown to detect by IKI? Can I see a loss of
>starch by staining seeds, or do I need to look at breakdown of starch in
>petri dishes? What concentration of GA will stimulate non-embryo seed
>halves to produce amylase? etc etc etc.
>
>Thanks in advance to anyone who responds!
>
>As an aside, I wonder how most schools respond to student such as this one
>- we had quite a debate about her in our department meeting! If your
>department has a policy for "conscientious objectors" in the lab classes
>I'd like to hear about it,

I asked about the Half-seed assay last fall (there is a thread of postings
last 11/25).  The consensus was 2% potato starch, 1% bacto-agar.  GA 10-4
-> 10-8 plus control.  I followed these suggestions.  Incubated plates 4
days, RT, in a drawer in the dark.  I then put them in the fridge the 3
days before class.  What we found was that at that concentration of starch,
the starch precipitated out.  Thus, we could visualize the halos WITHOUT
the KI (I subsequently stained to confirm for myself we were really seeing
satrch degradation).  Interestingly, the halo started getting smaller only
at 10-8.  On the other hand, big halos were seen around all others EXCEPT
10-4.  My guess is that this is a result of a dose response curve or such.
The students were impressed to see that.  A shorter incubation might have
better resolved the difference between 10-5 -> 10-8.  

I also came up with another useful control.  Have the student work up a
mouthful of saliva and spit on the plate.  The results are impressive.  It
also provided a nice connection for the students to us, and plants, food,
etc.  They were impressed alpha-maylase in our mouths worked on the plates.
 It may be a nice tie in for your student even though it isn't signalling,
etc.  By the way, I didn't use any antibiotics, etc. on the plates.  There
were a few threads of fungi growing but we were able to discount their
effect do to the patterns of halos versus fungal presence, hyphae, etc..

We have not had to deal with this particular issue yet, it hadn't even
occured to me as potential problem.  I have to admit that I avoided the
disection labs as an undergrad.  I always let my lab partner do the dirty
work, particularly when it came to working with anesthetised rats, pithed
frogs, etc.  As a TA I refused to take part in rat dissections though I did
teach the lab, ID parts for students, etc.  This caused a small disturbance
with the prof at first.  But my personal feelings were quickly accepted and
they worked around me, having another TA volunteer to come in and do the
demo cutting, etc.  This really wasn't based on any strong moral, ethical,
or religious belief, just a disdain for (what I consider to be) a needless
killing of animals for the sake of undergard bio majors.  Still, I was glad
to get the support of the prof and fellow TAs.

Our biggest conflict is with the teaching of evolution to our majors.  Some
are resistant enough to leave class when discussions lead to man being
related to ape, etc.  I often get disclaimers on exams stating something
like "I don't believe this crap, but I'll answer the questions as I think
YOU want them to be answered".


Dave Starrett


***********************************************************
* Dr. David Starrett                                      *
* Biology Department, MS 6200                             *
* Southeast Missouri State University                     *
* Cape Girardeau, MO  63701                               *
* Ph: 573-651-2382                                        *
* Fax: 573-651-2223                                       *
* Email: dstarret at biology.semo.edu                        *
* URL: http://biology.semo.edu/web/starrett/starrhpg.html *
*                                                         *
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