Extraction and STR Amplification of HAEIII Restriction Cut, Membrane Bound Human DNA

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dc.contributor.advisorJoseph Warren
dc.contributor.committeeMemberArthur Eisenberg
dc.contributor.committeeMemberJohn Planz
dc.creatorAndrews, John S.
dc.date.accessioned2019-08-22T21:35:01Z
dc.date.available2019-08-22T21:35:01Z
dc.date.issued2005-07-01
dc.date.submitted2014-05-01T06:48:05-07:00
dc.description.abstractThe use of DNA in forensics has become widely accepted since its introduction into the field in 1985 with Restriction Fragment Length Polymorphisms (RFLP) by Alec Jeffreys (1). RFLP techniques were utilized in the forensic DNA community until the mid 1990s when less labor-intensive PCR-STR techniques became available. During the transition from RFLP technology to PCR based STR technology a method for comparing RFLP profiles to that of STR profiles was not developed. Currently there have been no published studies where STR profiles have been obtained from membrane-bound, restriction cut human DNA. The only way to compare RFLP profiles to STR profiles would be to obtain STR profiles from the bound restriction cut DNA left on the nylon membranes. Since the shift in technology from RFLP to PCR-STR most labs, including the FBI, have stopped RFLP analysis as of the year 2000 (4). Today many unsolved cases exist that utilized RFLP technology. Due to the nature of RFLP analysis many times all of the biological sample must be consumed in order to obtain an RFLP profile. When this occurs, there is no longer biological sample left for future testing. In these instances the only DNA left from the case is restriction cut and bound to nylon RFLP membranes. The only chance of determining the STR profile of the source of the biological sample found at the crime scene would be to remove the membrane bound DNA and obtain an STR profile. The experimental hypothesis of this study is that DNA can be recovered from nylon membranes and interpretable STR results can be obtained. The use of multiple STRs are highly discriminatory being able to generate rare DNA profiles possessing a discriminatory power of in many times that of the earth’s population. Due to this discrimination power, profiles are able to individualize the source of a biological sample and aid in criminal investigations. If STR profiles could be obtained from old RFLP membranes numerous cold cases could be reopened and reinvestigated. The STR profiles obtained from the RFLP membranes could be placed into the Combined DNA Indexing System (CODIS). CODIS blends forensic science and computing software into a tool for solving violent crimes. Through CODIS, STR profiles can be entered into the database and searched against possible suspects at the local, state, and national level. Obtaining STR profiles from RFLP membranes would allow for the comparison of these profiles to those found in CODIS for a possible suspect. This project will employ methods to try and obtain an STR profile from Hae III restriction cut DNA bound to Magna Graph membranes. Attempts will be made to obtain STR profiles through direct amplification off of the membrane with PowerPlex 16 and separation on the Avant 3100 equipped with GeneMapper ID. Methods will also be utilized to remove the bound DNA from the membrane prior to amplification and separation. Removal of the bound DNA from the membrane will involve physical means, as well as, the use of various extraction chemicals. If a technique is found successful at removing DNA from Magna Graph membranes, then the technique will be applied to true RFLP membranes donated by the UNTHSC.
dc.format.mimetypeapplication/pdf
dc.identifier.urihttps://hdl.handle.net/20.500.12503/29439
dc.language.isoen
dc.provenance.legacyDownloads0
dc.subjectCell and Developmental Biology
dc.subjectCell Biology
dc.subjectCells
dc.subjectForensic Science and Technology
dc.subjectGenetics
dc.subjectGenetics and Genomics
dc.subjectGenetic Structures
dc.subjectLife Sciences
dc.subjectMedical Cell Biology
dc.subjectMedical Genetics
dc.subjectMedicine and Health Sciences
dc.subjectOther Cell and Developmental Biology
dc.subjectOther Genetics and Genomics
dc.subjectDNA
dc.subjectforensics
dc.subjectRFLP
dc.subjectPCR
dc.subjectSTR
dc.subjectbiological sample
dc.subjectmembrane bound DNA
dc.subjectSTR profile
dc.subjectCODIS
dc.subjectGeneMapper ID
dc.subjectPowerPlex 16
dc.titleExtraction and STR Amplification of HAEIII Restriction Cut, Membrane Bound Human DNA
dc.typeProfessional Report
dc.type.materialtext
thesis.degree.departmentGraduate School of Biomedical Sciences
thesis.degree.disciplineCell Biology and Genetics
thesis.degree.grantorUniversity of North Texas Health Science Center at Fort Worth
thesis.degree.nameMaster of Science

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