Investigation of Linear Amplification Using Abasic Site-Containing Primers Coupled to Routine STR Typing for LT-DNA...

Area of Science:

• Forensic Science
• Molecular Biology
• Genetics

Background:

• Obtaining complete short tandem repeat (STR) profiles from low template DNA (LT-DNA) is challenging due to stochastic effects and polymerase slippage.
• Existing amplification methods require improvement for accuracy and cost-effectiveness in LT-DNA analysis.

Purpose of the Study:

• To develop a novel, cost-effective, and accurate DNA amplification method for improving LT-DNA analysis.
• To address limitations in conventional methods for forensic STR typing.

Main Methods:

• Developed a novel linear amplification method (abLAFD) using primers with abasic sites near the 3' end to limit replication errors.
• Evaluated amplification efficiency using qPCR, showing a 130-fold increase in target DNA.
• Coupled abLAFD as a pre-PCR step with routine STR analysis (PP21) using 50 pg and 25 pg DNA.

Main Results:

• abLAFD demonstrated approximately a 130-fold increase in target DNA amplification.
• Coupling abLAFD with STR typing significantly improved allele recovery, average peak height, and heterozygote balance.
• The stutter ratio remained comparable to reference profiles, indicating method reliability.

Conclusions:

• The abLAFD method is a promising strategy for enhancing forensic LT-DNA analysis when coupled with STR typing.
• This technique offers a potential solution for obtaining more complete and reliable DNA profiles from minute DNA samples.