Jennifer M. Worley, Elaine S. Mansfield and Richard B. Rubin
Molecular Dynamics, 928 East Arques Avenue, Sunnyvale, CA 94086
DNA typing using multiplex short tandem repeat (STR) analysis is growing in popularity because of the method's exquisite sensitivity, speed, tolerance of degraded samples, and simplicity of typing small, discrete alleles. Both allelic ladders and standard molecular weight markers have been used as sizing standards for typing STRs. We have compared the accuracy of typing the Promega GenePrint™ Fluorescent Multiplex STR samples using an allelic ladder with typing using a variety of commercially available fluorescent sizing standards, with fragment intervals of 10 bp, 20 bp and 50 bp.
The samples were analyzed using the Molecular Dynamics FluorImager™ system. Alleles were called automatically using Genetic Typing software. By selecting appropriate loci from a table, this software may be used for typing D1S80 samples as well as monoplex and multiplex STR samples. Locus sizing tables may be edited to accommodate addition of new loci, adding new alleles for existing loci, and designing custom multiplex sets.
We found that locus-specific allelic ladders are consistently the most accurate sizing standards. Other molecular weight markers produced a 1.8 to 6.6 fold increase in sizing error for the largest locus, and a 1.2 to 14 fold increase in error for the smallest locus. This sizing error sometimes resulted in miscalled alleles. This increased error was observed in all the non-allelic ladders, regardless of fragment spacing interval in the markers or the number of sample lanes between markers. Though these markers might still allow accurate comparison of samples within a gel, it could incorporate errors in databasing of alleles.
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