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[Drosophila] overseeding

Leif Søndergaard via dros%40net.bio.net (by lsunicph from bio.ku.dk)
Fri Nov 16 03:08:46 EST 2012

Dear Mark,

I am pretty sure that your problem comes from what we in our lab call "crumble infection". It is an infection of wild yeast or another fungus which dries out the medium.
You can save infected cultures, at least partly, by adding a few drops of water.
To get rid of the infection you have to transfer flies to fresh vials a dozen times or so with one-two hour intervals.
Sometimes we seed our vials with dry bakers yeast (=live yeast) just before we use them and we rarely see "crumble infection" in such vials. But if this is a universal cure we have not investigated.

May all your Dros-experiments be succesful!

Best regards Leif Sondergaard
Dept og Biology; Univ. og Copenhagen

-----Original Message-----
From: dros-bounces from oat.bio.indiana.edu [mailto:dros-bounces from oat.bio.indiana.edu] On Behalf Of Mark Thorson
Sent: 15. november 2012 22:28
To: bionet-drosophila from magpie.bio.indiana.edu
Subject: Re: [Drosophila] overseeding

Laurence Mueller wrote:
> Mark,
> It is actually impossible to diagnose your particular problem with the
> information you have supplied. Female fecundity, and the resulting larval
> viability will be a function of (i) the age of the females, (ii) the size of
> the females (which depends on the larval density they were raised at), (iii)

The adults are about 0-3 days from emergence when
I use them to seed a new culture.  They look "normal"
size to me.  I see many wild flies every day (which
infiltrate my house through unscreened windows), and
they can be both larger and smaller than my flies,
although usually they're about the same size.

> the nutritional status of the females (e.g. were they well fed prior to
> laying eggs, where well fed means given excess live yeast), (iv) were they
> kept in a vials or bottles for several days prior to egg laying (the

They go directly from an old culture (14 to 17 days old)
to a new culture.  Old cultures do not seem to sustain
adults well, so I try to harvest soon after adult
emergence.  There usually are many larvae present in
an old culture.

> presence of many larvae in the female's environment will cause them to
> reduce egg laying and once they are given fresh food there will typically be
> an explosion of egg laying), (v) the volume and quality of the food provided
> for the larvae, (vi) the number of females laying eggs, and (vii) how long
> you let the females lay eggs (if you have lots of eggs in a culture there is
> no point to leaving adults in longer than one day since any eggs laid on
> days 2, 3 etc will be buried and killed by the feeding larvae that hatch
> from the first days egg laying).

I usually leave the adults in the cultures.  When I've
been short on adults, sometimes I would recycle them
from older cultures but not very often.  I have been
wondering whether leaving the adults in the cultures
is a bad practice, because they will lay eggs that will
never become adults.  I am concerned younger larvae
that hatch too late to pupate in my cultures might
contribute to stress on the older larvae which can
pupate.  If late eggs simply die, then I suppose
I should not worry about that possibility.  I have
been concerned about the waste of adults used for
seeding new cultures, but I haven't made much effort
to rescue them.  It's difficult to remove flies from
young cultures because the culture medium is not very
firm at that point, so I risk the medium spilling
if I tip the culture to dump out the adults.

> Finally it matters what your ultimate goal is. Do you want the largest
> number of flies possible, even if they are small, or do you want many, very
> large and therefore fecund flies. What's a worse outcome for you, occasional
> cultures that have very few flies but others that have many flies or having
> many cultures all with a moderate number of flies?

A large total number of flies is the best.  I'm
growing them for their sepiapterin content, so
ultimately total sepiapterin is the goal.

> It should be relatively easy to control the output of your cultures by
> standardizing the conditions I mentioned above, and then varying them
> systematically to find a set of conditions that works best for your goals.

A big part of my problem is not having good control
over several parameters, especially environmental
conditions, so I'm using a lot of intuition and
guesswork.  I'm thinking about turning a closet
into an incubator so I can at least get consistent
temperature control.  The recent cooler weather
seems to have greatly lengthened the life cycle.

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