Mutation Explorer™ Quick Users Guide Installation: Click the setup file of Mutation Explorer, the software will be installed to C:\program files\mutation Explorer\ unless you specify the differently. Open sample data at the directory of C:\program files\mutation Explorer\hetero…. SoftGenetics Inc. State College, PA 16803 Phone: 814-234-8483 Email: MutationExplorer@att.net www.softgenetics.com

Open Gene Information file 1. Click this fold will lead you to open file dialog box. 2. Click here will ask you to go to the directory to open the text file. You may go to c:\program files\mutation Explorer\hetero\gene_ infor.txt or select your own data for importation 3. Click “Add” will allow to input Genbank file *.seq by inputting c:\program files\mutation Explorer\hetero\*.seq. Input ab1 or scf sequence trace files to reference and sample areas of dialogue box in a similar fashion.

Run the program 4a. ME flow-chart (data input, contig, mutation calculation, report, and restart). You may click one by one or click report to get final results. 4b. Click “mutation report output” and analysis is completely automated finishing with Mutation Report in seconds

Mutation Report 5. Mutation Report: Red color, lower score; blue color, high score greater surety of mutation call. 6. Click any text of the report will lead you to the electropherogram, to confirm mutation call

Mutation Explorer Layout 7. Sequence texts of ref, sample, genbank. * means inconsistency 8. Amino acid sequence from genbank Reference electropherogram File manage-ment Sample electropherogram Genbank 3’5’ 9. Mutation Electropherogram. Mutation if it is a peak. Indel if center line turns red. Mutation peak table

Hot Keys Click the mutation peak table to zoom-into the mutation peak Hold Ctrl and click left mouse to manually select genbank, reference, or a mutation peak Draw a box from left to right with left mouse to zoom-in an electropherogram Draw a box from right to left to zoom-out. Hold left mouse and drag to move across electropherogram Trace quality boundary Click the mutation peak table to zoom-into the mutation peak

Mutation Parameters Intensity: mutation peak height. Area: the area of the mutation peak (x1000). Dropping factor: The percentage of intensity drop from reference to sample with respect to neighbor bases. Overlapping factor: the mutation peak relative to reference and sample peaks. Sn-ratio: signal-to-noise ratio. Mutation Score: accuracy score in logarithm scale. Comments: user may add comments.

Click the icon to change the red peak thresholds ME parameter settings Click the icon to change the red peak thresholds