Increasing Allele Detection by Altering the Quantity of Internal Lane Standard

Abstract

Electrokinetic injection (EI) is the primary method used in forensic laboratories to load amplified PCR product in capillary electrophoresis for short tandem repeat (STR) fragment separation. Because all samples subjected to capillary electrophoresis use internal lane standard (ILS), this study investigated the consequence of varying the volume of ILS and its effects on allele peak heights and number of alleles detected. Results demonstrated that when the volume of ILS is reduced, the average peak height and number of alleles increased, thereby increasing the sensitivity of the detection method. Sizing anomalies were observed; however, they did not adversely affect accuracy and precision. The method developed in this study offers a simple and universal procedure to increase the alleles detected in forensic STR analysis. Reducing the volume of ILS to achieve greater sensitivity is applicable to all STR amplification kits and capillary electrophoresis instruments currently used in forensic DNA analysis.

Automated detection and removal of capillary electrophoresis artifacts due to spectral overlap

“…Injection voltage was not included as a candidate variable because only single injection voltages were used for the 31XX series and 3500 series sample injections. Injection voltage will influence the total amount of DNA that is injected and therefore the peak heights, though increasing the voltage can also preferentially inject smaller fragments such as the smaller loci and primers . It is therefore possible that injection voltage could impact model performances, although this impact may be minimal due to the high level of information content within the currently used set of candidate variables.…”

“…The data were amassed from seven laboratories and included data run on 3 Applied Biosystems 3130 Generic Analyzers and 5 Applied Biosystems 3500 series Genetic Analyzers. 3500-run samples were injected for 15, 20, and 24s at 1 kV and 3130-run samples were injected for 3,5,10,11,12,15,20, and 30s at 3kV. Further information describing the sample set can be found in Appendix A in Supporting Information- Table 1A.…”