A Systemic Study Of Whole Genome Amplification Technology And Its Applications In Forensic Science

Objective : Genetic analysis from forensic microsamples is a urgent, difficult task in forensic science, because it is frequently limited by the amount of specimen available in forensic practice. Much effort has been carried out to resolve this difficulty. Whole genome amplification (WGA) technology has been thought to be a powerful, reliable and efficient strategy in analysis of minute amount DNA on many fields, for it can generate large quantity of DNA from minute DNA templates with unbiased way and its products can theoretically cover almost all the regions of genome. So, we employed two most used WGA methods-primer extension preamplification (PEP) and degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR) into forensic DNA analysis, and made a systemic study to evaluate its application in forensic science.Methods: We launch our study with the objective to get STRgenotyping result without STR slippage and no-specific products. We first observed factor that might result in STR slippage, and with these knowledge established standard and steady PEP and DOP approaches by optimizing several factors, then these approaches were used in single STR, multiplex STR loci, single shed hair and fingerprint DNA analysis for forensic purpose.Result: We found some factors that may result in STR slippage and even built up a STR slippage model by controlling several factors. The PEP and DOP approaches were established by optimizing several factors and proved to be reliable, unbiased, steady, sensitive, and efficient in forensic single STR analysis, multiplex STR analysis and single shed hair analysis. However, we did not succeed in STR genotyping of fingerprint sample, which may result from the DNA extracting method we used not sensitive enough. In silver detecting system, PEP and DOP methods can analys the quantity of DNA template at least 0.125ng in all samples, while the least needed quantity of DNA template in general PCR is 0.5ng. We also established an accurate, reliable method to get single cell through micromanipulation for DNA sensitive analysis and used it in our study. In multiplex PCR-ABI310 detecting system, PEP and DOP methods can succeed in genotyping Amelogenin and 9 loci in all studied samples at least 5 cells, while general PCR needed more than 10 cells.Conclusion: Approaches we have established in this study were reliable, steady enough for forensic purpose and can be used in forensic practice. The micromanipulation approaches we establishedhere was reliable and accurate method for DNA sensitive analysis. The PEP and DOP methods not only were more sensitive than general PCR, but also can produce large quantity DNA from microsample DNA, which can meet the need of both re-typing and check-typing for forensic purpose.