Modification of a commercially available kit for the improvement of PCR efficiency.
The polymorphism of short tandem repeats (STRs) is commonly used for human identity testing. Many commercial kits are available for this purpose, in which multiplex PCR for 10-16 STRs is usually used. For optimal results, the kits recommend rather limited conditions, which not all forensic samples can satisfy. We increased the efficiency of several commercial kits by modifying the components and reaction parameters (primer concentration and cycle number, and annealing and extension time). To simulate low copy number, we used 1:10 and 1:100 diluted samples compared to the recommended concentration. A change in cycle number, annealing and extension time, and primer concentration showed various results. Fine-tuning of PCR conditions by combining the described changes, decreasing the primer concentration, and increasing the annealing and extension time together with increasing the cycling number dramatically increased the efficiency of the reaction, even in low-copy-number simulated samples. Detailed information for several kit components or kits with different components targeting difficult sample conditions, including low copy number, would be of great help in forensics.
Hua Zhang
Ai
A
Department of Forensic Medicine and Institute of Forensic Science, College of Medicine, Seoul National University, 28 Yongon-Dong, Chongno-Gu, Seoul 110-799, Korea.
Bum Seo
Seung
S
Yi
Jin A
JA
Yeon Kim
Hye
H
Deok Lee
Soong
S
D004247:DNA / Q000032:analysis*; D004586:Electrophoresis; D044707:Forensic Sciences / Q000379:methods / Q000639:trends; D005784:Gene Amplification; D006801:Humans; D007723:Korea; D018895:Microsatellite Repeats / Q000235:genetics*; D010865:Pilot Projects; D016133:Polymerase Chain Reaction / Q000191:economics / Q000295:instrumentation*; D011110:Polymorphism, Genetic; D011933:Reagent Kits, Diagnostic / Q000592:standards*; D015203:Reproducibility of Results
D011933:Reagent Kits, Diagnostic; D004247:DNA