Different methods to determine length heteroplasmy within the mitochondrial control region

Abstract

The first and second hypervariable regions of the human mitochondrial DNA control region contain two homopolymeric stretches of cytosine (nt 16184-16193 and nt303-315, respectively). According to the Cambridge reference sequence these homopolymeric stretches are interrupted by thymine (T), at positions 16189 and 310, respectively. Monotonous runs of the same base have been suggested to be hot spots for mutations, probably caused by replication slippage, resulting in length heteroplasmy. This paper describes a rapid method based on restriction cleavage of labelled PCR products encompassing the homopolymeric tract in HVII to quantify the relative proportions of different length variants present in an individual. To compare the accuracy of this method, cloned PCR products from several heteroplasmic individuals have been additionally sequenced.

Mitochondrial DNA CA dinucleotide repeats in Koreans: the presence of length heteroplasmy

“…According to previous reports [15,16,17], direct sequencing, fluorescence-labelled restriction fragment analysis and cloning analysis have been used to detect and identify length heteroplasmy in mitochondrial HV1 and HV2 regions. In comparing the detection limit of the three methods, the level of detection was found to increase in the order from direct sequencing of the mtDNA to restriction fragment analysis, to the cloning assay [17]. However, restriction fragment analysis was thought to be a sufficiently accurate alternative for most applications involving heteroplasmy analysis.…”

Low-volume amplification on chemically structured chips using the PowerPlex16 DNA amplification kit

“…DNA analysis has become an indispensable tool in almost every forensic laboratory and is an integral part of main forensic tasks (i.e., paternity testing, human and nonhuman identification, and examination of forensic stains) [1][2][3][4][5][6]. The application of polymerase-based amplification of DNA-and especially of autosomal short tandem repeats (STRs)-to forensic molecular genetics has been pathbreaking [7][8][9][10].…”

Identification of Human Bone Remains by Autosomal STRs and Mitochondrial DNA SNPs

“…This finding agreed with that of several previous studies [23][24][25] . C-stretch length heteroplasmy increases the difficulties of DNA sequencing 19,20,36) and individual identification.…”

“…The evolutionary rate or base substitutions of both regions were shown to be high 17,18) . There is a C-rich sequence in both regions with the first being extensively studied [19][20][21] and its evolutionary mechanism is not certainly known 21) .…”

“…Moreover, there is a C-continuous tract or C-stretch in the HVS-I which is located in 16184 ~ 16193 nt 19) with a reference pattern 5C1T4C 28) . This tract acquires sequence-length variations and fast evolving among unrelated people [19][20][21] . The mechanism by which the C-stretch sequences were evolved is not certainly known.…”