Size of DNA Molecules Determined by Agarose Gel Electrophoresis

Dr. Chuck Kim, Facilitator

 

Physical Science Department

Cherry Hill High School East

1750 Kresson Rd

Cherry Hill, NJ 08003

856-424-2222 X222/332

Portrait by Abby Lichterman, Cherry Hill High School East, 1999

The distance DNA molecules can travel through agarose gel depends on the size of DNA molecules in electrophoresis. The next picture shows an example of gel electrophoresis.

Sample agarose gel electrophoresis DNA fragments

  In order to determine the size of DNA molecules, we need to compile data of the molecular size as a function of the migration distance and make a graph using a semi-log scale for y-axis, the molecular size, of a standard DNA such One Kilobase Ladder, as follow:   

Graph

 

 

 

 

 

 

 

 

 

 

 

 

This graph was produced by Microsoft Excel taking the following steps:

  1. Open Microsoft Excel.

  2. Enter all the data, keeping the migration distance in the first column and the size of molecules in the second column, as shown by the above example.

  3. Select the entire data and go to Chart Wizard.

  4. Select Chart Wizard > Chart Type-X-Y Scatter, and choose the graph with data points but not the ones with graph lines.

  5. Click Next and select Column in Data Ranges option. (Be patient and complete all five steps without skipping any options. Look into every option tab.)

  6. Click Next and enter all the information.

  7. In the Gridline option, enable both Major and Minor Gridlines.

  8. Click Finish to produce your graph.

  9. In order change the vertical scale to logarithmic scale, right-click on the vertical scale numbers on your graph.

  10. Choose Format Axis, open Scale tab, and enable Logarithmic. You can adjust the Minor Unit to 10 to produce enough grid lines.

  11. Right-click again on the X-axis scale to change the Minor Unit to 0.1 cm or 1 mm if you use mm as the minor unit in your data table.


Curve-Fitting:

 

 

 

 

 

 

 

After making your graph, you need make an equation that shows the relationship between the molecular size and the migration distance. 

  1. Do not select the graph but right-click one of the data point on your graph. This will open shortcuts for curve-fitting.

  2. Select Add Trendlines.

  3. Choose Power function as your Type. (Power function is really "powerful" because it can predict the most probable mathematical relationship.)

  4. Click Options tab and enable both Display Equation on Chart and Display R-Squared Value on Chart.

  5. Finally click OK to see you Equation and R-Squared value 


This type of curve-fitting is a powerful tool that our students can use in any scientific investigation. For more examples, go to Mathematical Modeling in Biology.