The analyst selects one of four possible size calling methods (in the analysis parameters) utilized by GeneScan^®:

1. Local Southern
2. 2nd (or 3rd) Order Least Squares
3. Cubic Spline
4. Global Southern method

The Local Southern method for size calling is the most commonly used algorithm in forensic DNA analysis. It determines the sizes of fragments by using the reciprocal relationship between fragment length and mobility. The unknown fragment is surrounded by two known sized fragments above and one below then two below and one above. The results are averaged and the size of the allele is determined.

For a review of size calling methods 2-4 listed above, reference the GeneMapper^® ID software version 3.1 User's Guide.⁰²

Below is a visual representation of the sizing process of the Local Southern method.

In variable-temperature environments, some studies have found that the Global Southern method (rather than the Local Southern method), may provide better sizing precision.^03, 04

One important issue concerning the sizing of AmpFℓSTR^® GS500 size standards (ROX or LIZ) is the de-labeling of the 250 base pair fragment. At a minimum, the 250 fragment should not be labeled.⁰⁴

Read more about delabeling in this course.

The size calling method is one component of the overall parameter settings that are configured prior to analyzing data.

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