0. GMO Teaching Supplement for DNA Extraction
A photo-guide with useful pointers
& tips for a successful DNA extraction
Contains photos documenting the key
steps in the DNA extraction procedure
Photos are numbered according to the
BABEC GMO Student & Teacher Guides
Can be used for a pre-lab discussion or
during lab for troubleshooting & support

1. 5. Internet Exploration:
Overview of GMO PCR Lab
1. Collect food or plant product
2. Isolate food/plant DNA
3. PCR of Plant and
35s or Bt genes
4. Gel
Electrophoresis
GMO-
GMO+
5. Internet Exploration:
“All About GMOs”
Fall 2011, page 1

2. Part 2: Removing Impurities
Lab Flow, Parts 1 & 2
Part 1: Food Lysis
Add plant or
food product
Add Lysis Buffer
Grind
Add Lysis Buffer
Mix
Boil
Part 2: Removing Impurities
Transfer 400l
of liquid
Add 40l
of NaCl
Centrifuge
Incubate on ice
Centrifuge
Fall 2011, page 2

3. Part 3: Isolating the DNA
Lab Flow, Part 3
Part 3: Isolating the DNA
Transfer 300l
of liquid
Add 400l of
Isopropanol
Mix
Centrifuge
Remove
liquid
Resuspend DNA
in TE/RNase
Dry DNA pellet
Store on ice
Fall 2011, page 3

4. Use a Small Amount of Food Material
The portion of food/plant material should sit
about halfway to the 0.1ml mark
(step 2)
0.1ml mark
The correct amount of corn meal
Fall 2011, page 4

5. Food Maceration Add 200l Lysis Buffer to food material (step 3)
Crush with micropestle
(step 4)
Twist and grind with an up & down motion
Keep food debris between pestle and tube wall
If necessary, remove stuck portion from
bottom of tube with pipette tip
Add 800l Lysis Buffer
to crushed material
(step 5)
Solid material may still remain,
but liquid should become cloudy
Fall 2011, page 5

6. Ideal Centrifuging Method
Orient the hinge of the tube to point outward and away from the middle of the centrifuge.
This allows for solid material to settle to the bottom of the tube directly below the hinge.
Hinge of tube points out
All spins should be performed at 10,000 x g,
which is about 10,000 rpm depending on the centrifuge
If you have a less powerful centrifuge,
spin longer than 5 minutes
below hinge
side view
After centrifuging,
look below the hinge for
the solid material (pellet)
Fall 2011, page 6

7. Removal of Food Lysate
Lysate is the solution produced when cells are destroyed.
You want to take the clear liquid, which contains the DNA, not the insect debris.
After centrifuging, there may be food debris
on bottom of the tube, and possibly on the top
Place your pipette tip between the two
layers and take 400l of the clear liquid
(step 9)
Lipids float
on top
Take liquid
from here
Proteins
settle to bottom
Fall 2011, page 7

8. Incubation on Ice The NaCl precipitates the SDS in the lysis buffer.
SDS can degrade DNA so you want to remove it.
Food lysate (400l)
with NaCl added (40l)
(step 10)
After sitting on ice, the NaCl
solution may become cloudy
because a precipitate has formed
ice
Fall 2011, page 8

9. Isolating the DNA
After the SDS precipitates out, the DNA is in the supernatant, the liquid portion.
After you centrifuge, the solid
material settles to the bottom.
Transfer 300l of the liquid into a new
tube and add 400l isopropanol
(step 13 & 14)
DNA is in the liquid
SDS; discard
Fall 2011, page 9

10. Drying the DNA Pellet
After the final centrifuge spin, the DNA is now in the bottom
of the tube because it precipitates with isopropanol
Pour off the liquid
& dry the pellet
(steps 16 & 17)
You may or may not see a pellet.
DNA is still there even if you can’t see anything.
Here are some examples of what you may see:
No visible DNA pellet
Small DNA pellet
Large DNA pellet
Your DNA is here,
directly under the hinge
Fall 2011, page 10

11. Resuspending the DNA
Depending on how large your pellet was, your DNA may be clear or cloudy.
DNA is resuspended in
200l TE/RNase
(step 20)
If DNA is cloudy or if there is visible debris,
add another 200l (step 21)
Cloudy DNA
DNA with debris
Clear DNA
Excessive food debris can inhibit the
PCR and it helps to dilute it out.
You should still have plenty of DNA.
Fall 2011, page 11