بهينه سازي روش I-PEP و تاثير آن بر نتايج حاصل از تعيين طرح واره با استفاده از تعداد اندك نسخه هاي DNA

Improvement in the I-PEP method and its effect on the outcome of low copy number DNA profiling

Background: DNA Profiling has become one of the most robust and reliable methods at forensic identification; However, an insufficient DNA quantity (less than 100 Pg or 33 copies) found often in forensic evidence samples, is a major hindrance. Amplification of such low copy number DNA samples is attainable with the most efficient whole genome amplification (WGA) method, named improved primer extension preamplification (I-PEP) PCR. Methods: By initial assessment of existing PEP and I-PEP methods on serially diluted DNA, it was attempted to reach an improved method leading to reliable profiling with the lowest amount of template. This method employs degenerate 15-mer PCR primers followed by specific amplification of DNA with specific primers. Findings: Subsequent to the amplification with the new modified and improved I-PEP, which we term KI-PEP PCR, complete DNA profile was obtained from only 2.5 pg of input DNA. Using this method, a fragment size of 1106 bp was effortlessly amplified with the specific primers. Conclusions: The Utility of KI-PEP PCR not only increases the low quantity of DNA, but also provides the optimum length appropriate to DNA Typing techniques.