Rapid amplification of the RM-Yplex assay

Area of Science:

• Forensic Genetics
• Molecular Biology
• DNA Analysis

Background:

• A previously developed multiplex PCR assay, RM-Yplex, targets 13 Rapidly Mutating Y STR loci.
• The original assay utilized Platinum® Taq DNA polymerase, requiring approximately 2.5 hours for amplification.

Purpose of the Study:

• To shorten the processing time of the RM-Yplex assay.
• To evaluate alternative DNA polymerases for faster and efficient amplification of Y STR loci.
• To maintain the reliability and robustness of the multiplex PCR assay.

Main Methods:

• Tested Phusion® Flash High Fidelity and TAKARA Z-taq™ DNA polymerases alongside Platinum® Taq DNA polymerase.
• Investigated various PCR cycling conditions for the RM-Yplex assay.
• Utilized Veriti® PCR thermal cycler for amplification.

Main Results:

• Achieved rapid, robust, and efficient amplification of all 13 Y STR markers within the multiplex.
• Reduced amplification time from 2.5 hours to less than 28 minutes using Phusion® Flash High Fidelity DNA polymerase.
• Confirmed reliable results with the optimized conditions.

Conclusions:

• Phusion® Flash High Fidelity DNA polymerase offers a significant time reduction for the RM-Yplex assay.
• The optimized assay provides a faster yet reliable method for Y STR analysis.
• This advancement improves efficiency in forensic genetic applications.