faculteit: "FNWI" en publicatiejaar: "2011"
| Auteur||Jorrit De Waele|
|Titel||Evaluation of factors affecting the identification of body fluids using mRNA in forensic case samples|
|Begeleiders||SallyAnn Harbison, Ate Kloosterman|
|Faculteit||Faculteit der Natuurwetenschappen, Wiskunde en Informatica|
|Opleiding||FNWI MSc Forensic science|
|Samenvatting||It is often important in forensic cases to determine the body fluid origin of a stain. Recently, ESR introduced CellTyper, a real-time PCR multiplex for a decisive identification of body fluids via mRNA profiling.
The aim of this thesis was the optimisation of the Promega Differex™ System process as part of the mRNA process. This kit for differentiation of sperm and epithelial cells did not work for mRNA analysis. The Digestion Solution was discovered to be the major factor imposing detriment to mRNA. Its Digestion Buffer component was found to degrade mRNA transcripts. Proteinase K appeared to contribute to this effect, but also weakened the spermatozoa leading to the release of sperm cellular material into the epithelial fraction and an incomplete differentiation. Replacement of the Digestion Solution by the Lysis Buffer from the Promega DNA IQ™ kit proved to promote mRNA stability, but could not achieve differentiation. The detrimental effect of the Digestion Buffer was counteracted by combination with the Lysis Buffer. This combined solution also gave phase separation. The optimal ratio of Lysis Buffer in Digestion Buffer is not yet established, but it was found to be less than 3/10. To date, differential extraction methods are not reliable for use with mRNA profiling in casework and it is recommended to perform a normal DNA IQ™ extraction instead.
mRNA profiling might replace presumptive test in the future, but until it is fully implemented in the forensic community, both methods will often be applied. Screening tests will also remain to be applied in cases when a quick indication is required. Therefore, it is important that presumptive testing does not impair mRNA profiling. This was also studied in this thesis.
Results obtained suggested that the Combur3 Test® for blood and the acid phosphatase test for seminal fluid do no infer with mRNA profiling, however it cannot be excluded that an effect would be detectable with very small stains. The Phadebas® treatment of saliva is likely to affect the mRNA, causing marker peaks to drop out or to be reduced. Careful considerations have to be made in this regard when testing.|
|Soort document|| scriptie master|
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