Downloading data from the FTP site
When sequencing results are ready we will send you an email notifying you that files are available to download from the FTP site. To gain access to your data, click onto Client FTP login and type in your Institute’s username and password (available from the facility). Files are retained on the FTP site for a period of three months and all data is backed up to permanent archive.
Sequencing files created.
Two file types are generated for each sample:
filename .ab1 file (raw data trace, electropherogram, annotation of the sample)
filename .seq file (text file of the sequence).
It is essential to check base calling accuracy using the electropherogram as reference. Manual editing especially at the end of the run may be required.
Programs to view sequence files:
Sequence scanner (Windows) is free viewing software provided by Applied Biosystems that enables you to view, edit, print and export sequence data and graphically express results in report format.
Chromas Lite (Windows) is freeware enabling you to view, print, export, copy and reverse/complement sequence data and search for sequences by exact matching or redundant codes.
FinchTV enables you to view chromatogram files on Mac OSX and Windows
4Peaks (Mac) enables you to view and edit sequence files without prior file conversion
Edit View (Mac) enables you to view chromatogram files
Macintosh users may need to convert the chromatogram from Win to Mac using the conversion utility from Applied Biosystems
Quality Values (QV) range from 1 to 50+ and are assigned to each base indicating the probability of error. The calculated QV is viewed using QV bars above each base call and can be viewed using the Applied Biosystems Sequence scanner program. The analysis sequencing program that the Centre uses assigns an “N” to any base with a QV < 15. pGEM and LongRead controls routinely generate 950 to 1000 QV>20 base pairs.
IUPAC/IUB Codes are assigned to mixed bases within the sequence. The code is defined by the International Union of Biochemistry.