What happens to your
Powdery Mildew
Samples once they arrive
in the lab?
To begin, I need to plant
some Baudin seed to use in The seed is planted in a
the lab.
substance
called
vermiculite, not soil.
The vermiculite is a
lightweight,
highly
absorbent mineral and
is used in thousands of
applications including
potting soils and grow
mixes.
The Baudin seeds are then given
water and left to grow under
lights in a temperature controlled
room. I give them additional
water every 2-3 days.
The lights in the room are on
a timer so that they are on
during the day and turn off at
night,
mimicking
the
conditions in the field.
While I'm waiting for the plants to
grow I need to prepare some agar
plates. Later, I will use the plates to
keep the different mildew samples
separate from each other in the
laboratory. I measure out some agar
powder and add it to water.
The agar must be sterilised before
use to remove bacteria and other
fungi. It is placed in an oven
where it is heated to 120°C for
20 minutes and then poured into
petri dishes. As the agar cools it
forms a gel.
Now I can start using your samples.
I cut leaf pieces from the Baudin I have
grown and place them at the bottom of
a plastic pipe.
I take your sample from the tube and
shake it over the top of the pipe.
Spores of the fungi move from your
sample onto my fresh Baudin leaves.
The leaves are then inserted into the
agar gel in the petri dishes. The
dishes are sealed with tape to keep
the isolates separate from one
another.
They are then left to grow.
In about 7 to 10 days, the
fluffy mildew begins to appear
on the infected leaves.
Now I can start to get DNA
from the isolates.
Using the plates I prepared
earlier, I cut away a single
fluffy pustule and place it into
a small tube.
To process the swabs I cut
off a small piece of the
cotton tip and place it into
a separate tube.
I use my pipette to add
several different
liquids into the tubes.
A
pipette
is
an
instrument used to
measure out very small
volumes of liquids.
The tubes are then put into a
machine which heats and cools
the liquids in a programmed
order. This process causes a
reaction which greatly
increases the amount of DNA
in each tube.
Once the reaction is complete, I use a
method called gel electrophoresis to
look at the quality and quantity of my
DNA.
I use my pipette to suck
up a tiny amount of DNA
and load it into small
wells in a gel.
The gel is in a tank filled with a special
liquid. After loading, the tank is subject to
an electric current which causes the DNA to
move down through the gel. The pieces of
DNA move according to their length. Longer
pieces move slower, smaller pieces move
faster.
After about 60 minutes the gel is ready to use.
I take the gel to a special machine which
shines an ultra violet light through the gel.
The machine puts out a picture which
allows me to quantify and qualify the
DNA.
Afterwards, I submit the DNA for
sequencing.
This allows me to check for mutations
which cause resistance towards some
fungicides.
This is an example of the DNA
sequence from one of the
isolates.
So that’s what happens
to your Powdery
Mildew Samples once
they arrive in the lab.
Thanks so much for
your contribution to my
work.