ABI sequencing analysis |
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Authors: | Tracy L Hagemann Sau-Ping Kwan |
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Institution: | (1) Department of Pathological Sciences, The Waisman Center, University of Wisconsin, 53706 Madison, WI |
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Abstract: | The ABI Sequencing Analysis application is designed specifically for the analysis of data produced by the ABI DNA Sequencer.
The ABI sequencer is a laser-based instrument that utilizes fluorescent labels to analyze the products of a sequencing reaction
as they migrate through a gel. After the data are collected from a sequencing run, the Analysis program identifies and tracks
the sample lanes of the gel and subsequently normalizes and integrates the raw data into a chromatogram of the final sequence.
For the use, there are basically two types of files that can be manipulated to potentially improve the analysis results. The
Gel File consists of a computer generated image of the sequencing gel with the fluorescent DNA banding patterns. This image
allows the user to view and edit the tracking lines generated and used by Analysis to collect data points for each sample.
Individual Sample Files are stored for each of the samples analyzed and include the chromatogram, raw data, and annotations
and information regarding the sample and sequence run. Generally, the products of a sequencing reaction are easily resolved
and the Analysis software interprets the correct nucleotide sequence. Ambiguous base calls tend to occur near the end of the
sequence and may be either edited or deleted by the user before exporting the data for further comparisons or alignments.
Occasionally the tracking lines within the gel image may need to be adjusted or moved. The sample data are then reextracted
from the Gel File and analyzed again. This review explains the general operation of Analysis in terms of viewing and editing
a chromatogram, retracking the lanes of a Gel File, and analyzing the final sample data. The three versions 1.2.1, 2.1.2,
and 3.3 are discussed. |
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Keywords: | Chromatogram gel file sample file tracking sample manager basecaller |
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