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Amplified Fragment Length Polymorphism (AFLP) Analysis on Applied Biosystems Capillary Electrophoresis Systems

ysis with a specific application, a parentage assay was performed on maize DNA samples isolated from three individuals:

1. Parent one (P1)

2. Parent two (P2)

3. The first-generation offspring (F1) from a cross between P1 and P2

Primers labeled with the 5-FAM dye from the Applied Biosystems AFLP starter kit were used for selective amplification, and the reaction products were analyzed using a 36-cm capillary array and POP-7 polymer. The reaction products were mixed with the GeneScan-500 ROX dye-labeled size standard and electrophoresed on the 3130xl Genetic Analyzer and the 3730xl DNA Analyzer using the following protocols:

FA_36_POP-7 run module and F dye set (3130xl Genetic Analyzer)

GeneMapper_36_POP-7 run module and Any4Dye set (3730xl DNA Analyzer)

For the 3730 series systems the user should review all the data from the spectral calibration when using the AnyDye feature to verify that no poor quality data was used. Condition Numbers and Quality Score boundaries should be optimized for each dye set. Refer to the BAC Fingerprinting on the Applied Biosystems 3730/3730xl DNA Analyzer application note (S/N 107AP04-01) for more detail on this procedure.

The peak patterns in the F1 sample are consistent with those of a cross between P1 and P2 (Figure 3). In addition to several non-polymorphic peaks in both P1 and P2, which are also present in F1 (black arrow in Figure 3), unique peaks present in the F1 sample that were inherited from one parent (red arrow in Figure 3) can also be identified. Researchers can use the custom plot colors option in GeneMapper software for easy identification of polymorphic peaks (red arrow) and common peaks (black arrow). For example, in the data shown in Figure 4, a polymorphic peak can be easily identified in the P1 sample (red arrow). The use of the POP-7 polymer offers advantages such as high resolution, especiall
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