HUMAN D-LOOP mtDNA MUTATIONS OF BLOOD CELLS, EPITHEL CELLS, AND HAIR FOLLICLES FROM CERTAIN INDIVIDUAL WITH DIFFERENT AGES

Abstract: <br /> <br /> <br /> <br /> <br /> One of the unique characteristics of mtDNA is its mutation rate which is relatively higher than the DNA nucleus. The high mutation rate of mtDNA causes high level of polymorphism among individuals. In mtDNA, there is a n...

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Bibliographic Details
Main Author: (NIM : 204 05 018), RAifuddin
Format: Theses
Language:Indonesia
Online Access:https://digilib.itb.ac.id/gdl/view/6632
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Institution: Institut Teknologi Bandung
Language: Indonesia
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Summary:Abstract: <br /> <br /> <br /> <br /> <br /> One of the unique characteristics of mtDNA is its mutation rate which is relatively higher than the DNA nucleus. The high mutation rate of mtDNA causes high level of polymorphism among individuals. In mtDNA, there is a noncoding region known as displacement loop (D-loop) which has two hypervariable regions, I and II (HVR I and HVR II). However, there have not been any information whether the sequence of D-loop mtDNA nucleotides is similar either for different cells of certain individual or for individuals with different ages. This study is aimed at obtaining the information of human D-loop mtDNA mutation for different cells in each individual of five individuals with different ages. This study consists of several stages, namely preparation of DNA template, amplification, nucleotides sequencing, and mutation analysis. Preparation of mtDNA template employing cell lysis method, then followed by amplification of D-loop mtDNA using the Polymerase Chain Reaction (PCR) with M1 and HV2R primers, then the analysis of PCR result using agarose gel electrophoresis with the pUC19/HinfI as standard. Amplified DNA was sequenced using the dideoxy Sanger method with M1 and M2 primers. DNASTAR program was used to analyzed nucleotides mutation with the Cambridge Reference Sequence (CRS) method as standard. There were 12 different cells as samples (blood cells, epithel cells and hair follicles) which were originated from five individuals with different ages (10, 20, 30, 40, and 80 years-old respectively). PCR amplification results of all template DNA samples showed a single band of DNA fragment which was estimated as 1 kb length. The result of nucleotide sequence, in the form of electropherogram, indicated that the seven samples (from three individuals) and five samples (from two individuals) were 200 and 600 base pairs respectively, with the total of about 5.500 base pairs readable nucleotide sequence.The result of the nucleotide sequence analysis using CRS as the standard sequence indicated that the position and the type of mutation of the 30 and 40 years old individuals were similar for blood cells, epithel cells, and hair folicles. In addition, the analysis of different cells in 10, 20, and 80 years old individuals also indicated the similar position and type of nucleotide sequence mutation for blood cells and hair folicles in each of individual. <br /> <br /> <br /> <br /> <br /> This work demonstrated that the nucleotide sequence of D-loop mtDNA for different cells such as blood cells, epithel cells and hair folicles of each individual showed similar mutation. This is due to the same source of cell namely ovum. The ovum has one type of mtDNA which is differentiated align with the growth of embrio. This differentiation did not cause nucleotide sequence mutation of mtDNA in the blood cells, epithel cells, and hair follicles of an individual. It is suggested that for the mtDNA based forensic identification, single type of the above cells would be adequate and would contribute to the more simple identification process in forensic.