Académique Documents
Professionnel Documents
Culture Documents
MALAYSIAN INSTITUTE OF
CHEMICA L & BIOENGINEERING TECHNOLOGY
CBB 30204
BIOSEPA RATIONS ENGINE ERING
TECHNOLOGY
EXPERIMENT 2
CELL LYSIS
OBJECTIVES
1. isolate DNA from onion tissue.
2. make observations regarding the results of the isolation.
3. develop a hypothesis from their observations regarding the nature of the
molecule.
4. design an investigation to test their hypothesis
EXPERIMENTAL PROCEDURES
1. Prepare the homogenization solution ahead of time (no more than 24 hrs before). The
ethanol should be chilled overnight in a freezer, with the cap loosened.
Homogenization Medium (100 mL)
to 80 mL of distilled water add:
3 mL liquid dish soap (ex. Joy)
0.8 g NaCl
0.4 g sodium citrate
0.3 g EDTA
CBB 30204 Bioseparations Engineering Technology
9. Slowly layer the appropriate amount of chilled ethanol on top of the homogenate. A
layering effect can be achieved by tilting the beaker containing the cell homogenate and then
slowly pouring the ethanol down the side of the beaker. White strands of precipitated DNA
may appear.
10. Using a glass stirring rod, wind the DNA around the rod by slowly spinning the glass rod
while stirring the mixture in one direction. Strands of precipitated DNA should begin to
appear at the interface of the cell homogenate/ethanol mixture. Continue stirring and
collecting precipitated DNA for a few minutes.
11. Once the DNA is collected, gently scrape it into a 1.5 mL micro centrifuge tube that
contains 1 mL of water or low salt buffer (e.g. TE). If stored in a refrigerator, the DNA
sample can last for several months and be used for other experiments.
ASSIGNMENT
1. Determine and explain the principle(s) of cell lysis applied in the experiment.