2 Dimensional Electrophoresis - RITJBF/JBioFramework GitHub Wiki

The 2D electro1DMain simulation can accept input in three different file formats: FASTA, Genbank, and Protein Data Bank. Individual protein spots in the final output are linked to data about that protein and to external databases. In addition to the on-screen simulation of the separation process, the application also generates output in the form of HTML and spreadsheet files that contain sequence, function, molecular weight and pI values.

#Loading Proteins onto Gel

Clicking on the Add Proteins button will pop up a dialog box where you can open a protein file:

Protein files of either Fasta, Genbank, or PDB format can be added by simply selecting the appropriate file from the list as seen below and then clicking the Load button:

Once the file has finished loading and the simulation is ready to proceed, the following message box will appear:

#Animation Buttons

There are three animation buttons used for running the Electrophoresis gel; Run, Stop, and Restart. During an SDS-Page animation, the Run button will change to the Pause button.

The Run button will begin the selected animation.

The Run button will change to the Pause button when an SDS-Page animation is in progress; click it to freeze the gel in the condition it is in. If you wish to run the gel for a longer period of time after the animation has stopped, click the Run button again.

The Stop button will return the gel to the condition it was in before either the IEF or SDS-Page animations were run, but with the protein list(s) still loaded, along with any changes those lists may have undergone.

The Reset button will return all of the conditions of the gel back to what they were when the JBioFramework program was first opened. This includes unloading the protein lists and clearing the gel canvas.

#Separation conditions

Before running the separation it is important to specify the conditions (pH range and % acrylamide) on which to run the gel.

The drop down Choose PH menu will allow you to select the pH range you want to give the gel.

Additionally, you can specify a range by selecting "Enter A Range" and typing in a min and max pH.

There is also a drop down menu to allow you to select what percentage of Acrylamide to use.

#Isoelectric Focusing (IEF)

Once your proteins have been loaded, IEF should be displayed as the "Current Animation".

Pushing the run button in "Animation Buttons" will start the isoelectric focusing simulation similar to below.

Various pH ranges might look similar to this:

#SDS-PAGE

Once the Isoelectric focusing simulation has finished, the "Current Animation" will switch to "SDS-PAGE".

Pushing the run button in the "Animation Buttons" area will now start the SDS-PAGE simulation similar to below:

The simulation can be paused, stopped, or restarted using the Animation Buttons. Pushing the run button again after the simulation has automatically stopped will cause the gel to run for another period of time. Pushing the restart button will set the program back to the condition it was in when first started- with no protein files loaded.

#Exploring the Separation

#Display Protein List

#Search Proteins List

#Generate HTML

Clicking the Generate HTML button will open the Sort Web Page Frame.

This allows you to create a table-formatted HTML document of all the proteins currently loaded in the display. The .html file that is created is stored in the "HTML files" folder of JBioFramework.

Sorting the HTML document by "Protein Title" will sort the proteins by gi-accession number. Sorting by "pI Value" will sort the proteins from least to greatest pI value. Sorting by "Molecular Weight Value" will sort the proteins from least to greatest molecular weight. Finally, sorting by "Protein Function" will sort the proteins alphabetically based on the first word in the function description. Below is an example HTML document sorted by "Molecular Weight Value":

#Record to CSV

Clicking on the Record to CSV Button will appear to do nothing. However, it is creating a comma separated file of the essential values of each protein in the currently loaded set and placing it in the CSV Files folder.

The file can be imported into spreadsheet programs like Microsoft Excel for further analysis. The file contains the titles of the proteins, their amino acid sequences, molecular weights, pI values, and X and Y coordinates on the 2-D gel.

#Color Key

Click the Color Key button to bring up a color key which will help you in determining which protein dots on the gel canvas are what types of protein.

#Adding New Protein Files