Restriction Digestion and Agarose Gel Electrophoresis


Subject Area Biology
Age/Grade Level Freshmen or Sophomores
Estimated Length     2 50-minute class period
Prerequisite Knowledge/Skills Students should have a general understanding about DNA structure and function.
Description of
New Content		 This lesson introduces students to the topics of restriction digestion of DNA and to agarose gel electrophoresis.  Students will use both techniques to analyze a sample of DNA.
Goals Students will use restriction enzymes to cut lambda DNA into various sized fragments.  The students will then run the DNA samples on a gel to see different banding patterns that can be achieved with different enzymes. 
Materials Needed Three different restriction enzymes, lambda DNA, enzyme buffer, agarose, water, test tubes, a uv lamp,  micropipettors, gel boxes or the wardsci science restriction enzyme kit found here.  
Procedure
 Powerpoint here. Have the students label three tubes with the names of the enzymes that you will use.   The reaction setups are as follows:
sterile ddH20        q.s (where "q.s." means quantity sufficient)
10X assay buffer     one-tenth volume
DNA                  x ul
restriction enzyme*  y ul (1-10 units per ug DNA)
Total volume         z ul
The reaction should be set up during the first class period.  The second class period will be used for agarose gel electrophoresis.  To make the agarose gel, approximately 1 gram of agarose should be used per 100 mL of water.  The gels should be made before the class period in order to save time.  It is possible for the students to set the gels up during the first class period. Once the gels have set, load the wells using the micropippetors and place the gels in the gel boxes.  Run the electric current for a half hour.  Make sure the DNA runs towards the positive side.  Once the gel has run for a half hour, use the UV lamp in a dark area to see the banding patterns.  Have the students draw the banding patterns that they see.
Evaluation See worksheet here.
Extensions This lesson could lead into a lesson on DNA transformation or viruses.