scholarly journals Unusual mtDNA Control Region Length Heteroplasmy in the COS-7 Cell Line

Genes ◽  
2020 ◽  
Vol 11 (6) ◽  
pp. 607
Author(s):  
Nataliya Kozhukhar ◽  
Sunil Mitta ◽  
Mikhail F. Alexeyev
Keyword(s):  
Mitochondrial Genome ◽  
Cell Line ◽  
Control Region ◽  
Tandem Repeats ◽  
Variable Number ◽  
Length Heteroplasmy ◽  
Repeat Units ◽  
Chlorocebus Aethiops ◽  
Region Length ◽  
Transcription And Replication

The COS-7 cell line is a workhorse of virology research. To expand this cell line’s utility and to enable studies on mitochondrial DNA (mtDNA) transcription and replication, we determined the complete nucleotide sequence of its mitochondrial genome by Sanger sequencing. In contrast to other available mtDNA sequences from Chlorocebus aethiops, the mtDNA of the COS-7 cell line was found to contain a variable number of perfect copies of a 108 bp unit tandemly repeated in the control region. We established that COS-7 cells are heteroplasmic with at least two variants being present: with four and five repeat units. The analysis of the mitochondrial genome sequences from other primates revealed that tandem repeats are absent from examined mtDNA control regions of humans and great apes, but appear in lower primates, where they are present in a homoplasmic state. To our knowledge, this is the first report of mtDNA length heteroplasmy in primates.

Profound Length Heteroplasmic Alterations in Mitochondrial DNA Control Region From Primary AML Cells: Implication of Impaired Mitochondrial Replication and Demonstration of ‘vicious cycle’ Hypothesis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3116-3116
Author(s):  
Myung-Geun Shin ◽  
Hye Ran Kim ◽  
Hyeoung-Joon Kim ◽  
Hoon Kook ◽  
Tai Ju Hwang ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Control Region ◽  
Copy Number ◽  
Regulatory Region ◽  
Mtdna Control Region ◽  
Vicious Cycle ◽  
Mtdna Copy Number ◽  
Length Heteroplasmy ◽  
Region Length ◽  
D Loop

Abstract Abstract 3116 Poster Board III-53 Mitochondrial DNA (mtDNA) control region (displacement (D)-loop including HV1 and HV2) is a non-coding region of 1124 bp (nucleotide positions, np 16 024–576), which acts as a promoter for both the heavy and light strands of mtDNA, and contains essential transcription and replication elements (Blood 2004;103:4466-77). Importantly, mutations in the D-loop regulatory region might change mtDNA replication rate by modifying the binding affinity of significant trans-activating factors (Eur J Cancer 2004;40:2519-24). Thus, length heteroplasmic alterations of mtDNA control region may be related with mitochondrial dysfunction resulting in ‘vicious cycle’ (Mol Med Today 2000;6:425-32). In an attempt to investigate profiling of mtDNA length heteroplasmic alterations in primary AML cells, we carried out a quantitative size-based PCR product separation by capillary electrophoresis (ABI 3130XL Genetic Analyzer and ABI Prism Genotyper version 3.1) using six targets (np 303-315 poly C, np 16184-16193 poly C, np 514-511 CA repeats, np 3566-3572 poly C, np 12385-12391 poly C and np 12418-12426 poly A). Length heteroplasmy was further confirmed by cloning and sequencing. Quantitative analysis of mtDNA molecules was performed using the QuantiTect SYBR Green PCR kit (Qiagen) and Rotor-Gene 3000 (Corbett Research). Forty-eight AML bone marrow samples were collected after receiving Institutional Review Board approval and informed consent. There were profound alterations of mtGI in 303 poly C, 16184 poly C and 514 CA repeats. The length heteroplasmy pattern of 303 poly C tract in the HV2 region disclosed mixture of 7C, 8C, 9C and 10C mtDNA types. In the HV2 region, length heteroplasmy in poly-C tract at np 303 - 309 exhibited 5 variant peak patterns: 7CT6C+8CT6C (50.0%), 8CT6C+9CT6C (14.0%), 8CT6C+ 9CT6C+ 10CT6C (10.4%), 9CT6C+10CT6C+11CT6C (8.3%) 9CT6C + 10CT6C + 11CT6C+12CT6C (2.1%). The length heteroplasmy pattern of 514-523 CA repeats in the HV2 region exhibited 2 variant peak patterns: CACACACACA (56.3%) and CACACACA (43.7%). In the HV1 region, length heteroplasmy in the poly-C tract at np 16184 - 16193 exhibited 9 variant peak patterns: 5CT4C+5CT3C (31.0%), 6CT4C+6CT3C (2.1%), 9C+10C+11C+12C (16.7%), 9C+10C+11C (2.1%), T4CT4C+5CT3C (4.2%), 9C+10C+11C+12C+13C (2.1%), 3CTC4C+5CT3C (2.1%), 10C+11C+12C+13C (4.2%), 8C+9C+10+11C (2.1%). Primary AML cells revealed decreased enzyme activity in respiratory chain complex I, II and III. AML cells had about a two-fold decrease in mtDNA copy number compared with normal blood mononuclear cells. Current study demonstrates that profound length heteroplasmic alterations in mtDNA control region of primary AML cells may lead to impairment of mitochondrial biogenesis (reduction of mtDNA copy number) and derangement of mitochondrial ATP synthesis. During this perturbation, mitochondria in primary AML cells might produce a large amount of reactive oxygen species, which causes the vicious cycle observed in chronic inflammatory diseases and cancers as well. Disclosures No relevant conflicts of interest to declare.

scholarly journals Using high-resolution annotation of insect mitochondrial DNA to decipher tandem repeats in the control region

2018 ◽  
Author(s):  
Ji Haishuo ◽  
Xu Xiaofeng ◽  
Jin Xiufeng ◽  
Cheng Zhi ◽  
jin Hong ◽  
...  
Keyword(s):  
Steady State ◽  
High Resolution ◽  
Mitochondrial Genome ◽  
Control Region ◽  
Transcription Initiation ◽  
Tandem Repeats ◽  
Mitochondrial Gene ◽  
Mitochondrial Genomes ◽  
Small Rna Sequencing ◽  
Antisense Gene

In this study, we used a small RNA sequencing (sRNA-seq) based method to annotate the mitochondrial genome of the insect Erthesina fullo Thunberg at 1 bp resolution. Most of the new annotations were consistent with the previous annotations which were obtained using PacBio full-length transcripts. Two important findings are that animals transcribe both entire strands of mitochondrial genomes and the tandem repeat in the control region of the E. fullo mitochondrial genome contains the repeated Transcription Initiation Sites (TISs) of the H-strand. In addition, we found that the copy numbers of tandem repeats showed a great diversity within an individual, enriching the fundamental knowledge of mitochondrial biology. This sRNA-seq based method uses 5′ and 3′ end small RNAs to annotate nuclear non-coding and mitochondrial genes at 1 bp resolution and can also be used to identify new steady-state RNAs, particularly long non-coding RNAs (lncRNAs). Animal mitochondrial genomes containing one control region only encode two steady-state lncRNAs, which are the Mitochondrial D-loop 1 (MDL1) and its antisense gene (MDL1AS), while all other reported mitochondrial lncRNAs could be degraded fragments of transient RNAs or random breaks during experimental processing. The high-resolution annotations of mitochondrial genomes can be used to study the phylogenetics and molecular evolution of animals or to investigate mitochondrial gene transcription, RNA processing, RNA maturation and several other related topics.

scholarly journals Dynamic of Mutational Events in Variable Number Tandem Repeats ofEscherichia coliO157:H7

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
A. V. Bustamante ◽  
A. M. Sanso ◽  
D. O. Segura ◽  
A. E. Parma ◽  
P. M. A. Lucchesi
Keyword(s):  
Mutation Rate ◽  
Tandem Repeats ◽  
Serial Passage ◽  
Variable Number ◽  
Mutation Rates ◽  
Similar Proportion ◽  
Factors Affecting ◽  
Repeat Units ◽  
Differential Weighting ◽  
Epidemiology Studies

VNTRs regions have been successfully used for bacterial subtyping; however, the hypervariability in VNTR loci is problematic when trying to predict the relationships among isolates. Since few studies have examined the mutation rate of these markers, our aim was to estimate mutation rates of VNTRs specific for verotoxigenicE. coliO157:H7. The knowledge of VNTR mutational rates and the factors affecting them would make MLVA more effective for epidemiological or microbial forensic investigations. For this purpose, we analyzed nine loci performing parallel, serial passage experiments (PSPEs) on 9 O157:H7 strains. The combined 9 PSPE population rates for the 8 mutating loci ranged from 4.4 × 10−05to 1.8 × 10−03mutations/generation, and the combined 8-loci mutation rate was of 2.5 × 10−03mutations/generation. Mutations involved complete repeat units, with only one point mutation detected. A similar proportion between single and multiple repeat changes was detected. Of the 56 repeat mutations, 59% were insertions and 41% were deletions, and 72% of the mutation events corresponded to O157-10 locus. For alleles with up to 13 UR, a constant and low mutation rate was observed; meanwhile longer alleles were associated with higher and variable mutation rates. Our results are useful to interpret data from microevolution and population epidemiology studies and particularly point out that the inclusion or not of O157-10 locus or, alternatively, a differential weighting data according to the mutation rates of loci must be evaluated in relation with the objectives of the proposed study.

Complete mitochondrial genome of the endangered Mary River turtle (Elusor macrurus) and low mtDNA variation across the species' range

2016 ◽  
Vol 64 (2) ◽  
pp. 117 ◽  
Author(s):  
Daniel J. Schmidt ◽  
Brittany Brockett ◽  
Thomas Espinoza ◽  
Marilyn Connell ◽  
Jane M. Hughes
Keyword(s):  
Mitochondrial Genome ◽  
Control Region ◽  
Tandem Repeats ◽  
Complete Mitochondrial Genome ◽  
Illumina Miseq ◽  
Base Pairs ◽  
Pet Trade ◽  
Nucleotide Variability ◽  
Natural Range ◽  
Miseq Platform

Elusor macrurus is an endangered short-necked turtle restricted to the Mary River catchment in south-eastern Queensland. Shotgun sequencing of genomic DNA was used to generate a complete mitochondrial genome sequence for E. macrurus using the Illumina MiSeq platform. The mitogenome is 16 499 base pairs (bp) long with 37 genes arranged in the typical vertebrate order and a relatively short 918-bp control region, which does not feature extensive tandem repeats as observed in some turtles. Primers were designed to amplify a 1270-bp region that includes 81% of the typically hypervariable control region. Two haplotypes were detected in a sample of 22 wild-caught individuals from eight sites across its natural range. The Mary River turtle is a species with low mtDNA nucleotide variability relative to other Chelidae. The combination of a very restricted distribution and dramatic reduction in population size due to exploitation for the pet trade are the conditions likely to have led to very low mtDNA variability in this endangered species.

scholarly journals High Sequence Variations in Mitochondrial DNA Control Region among Worldwide Populations of Flathead MulletMugil cephalus

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Brian Wade Jamandre ◽  
Jean-Dominique Durand ◽  
Wann-Nian Tzeng
Keyword(s):  
Mitochondrial Dna ◽  
Control Region ◽  
Tandem Repeats ◽  
Evolutionary Rate ◽  
Variable Number ◽  
Rapid Radiation ◽  
Mitochondrial Dna Control Region ◽  
Good Marker ◽  
Sequence Variations ◽  
Complete Mtdna

The sequence and structure of the complete mtDNA control region (CR) ofM. cephalusfrom African, Pacific, and Atlantic populations are presented in this study to assess its usefulness in phylogeographic studies of this species. The mtDNA CR sequence variations amongM. cephaluspopulations largely exceeded intraspecific polymorphisms that are generally observed in other vertebrates. The length of CR sequence varied amongM. cephaluspopulations due to the presence of indels and variable number of tandem repeats at the3′hypervariable domain. The high evolutionary rate of the CR in this species probably originated from these mutations. However, no excessive homoplasic mutations were noticed. Finally, the star shaped tree inferred from the CR polymorphism stresses a rapid radiation worldwide, in this species. The CR still appears as a good marker for phylogeographic investigations and additional worldwide samples are warranted to further investigate the genetic structure and evolution inM. cephalus.

scholarly journals Development of a new nomenclature for Salmonella Typhimurium multilocus variable number of tandem repeats analysis (MLVA)

2009 ◽  
Vol 14 (15) ◽  
Author(s):  
J T Larsson ◽  
M Torpdahl ◽  
R F Petersen ◽  
G Sørensen ◽  
B A Lindstedt ◽  
...  
Keyword(s):  
Salmonella Typhimurium ◽  
Tandem Repeats ◽  
Fragment Size ◽  
Foodborne Pathogen ◽  
Variable Number ◽  
Size Analysis ◽  
Fragment Analysis ◽  
Repeat Units ◽  
Reference Strains ◽  
Selection Of

Multilocus variable number of tandem repeats analysis (MLVA) has recently become a widely used highly discriminatory molecular method for typing of the foodborne pathogen Salmonella Typhimurium. This method is based on amplification and fragment size analysis of five repeat loci. To be able to easily compare MLVA results between laboratories there is a need for a simple and definitive nomenclature for MLVA profiles. Based on MLVA results for all human S. Typhimurium isolates in Denmark from the last five years and sequence analysis of a selection of these isolates, we propose a MLVA nomenclature that indicates the actual number of repeat units in each locus. This nomenclature is independent of the equipment used for fragment analysis and, in principle, independent of the primers used. A set of reference strains is developed that can be used for easy normalisation of fragment sizes in each laboratory.

Tandem repeats and length variation in the mitochondrial DNA control region of Epirrita autumnata (Lepidoptera: Geometridae)

Genome ◽  
2002 ◽  
Vol 45 (5) ◽  
pp. 855-861 ◽  
Author(s):  
N Snäll ◽  
K Huoponen ◽  
M -L Savontaus ◽  
K Ruohomäki
Keyword(s):  
Mitochondrial Dna ◽  
Control Region ◽  
Tandem Repeats ◽  
Repetitive Sequences ◽  
Distinct Type ◽  
Variable Number ◽  
Tandem Array ◽  
Epirrita Autumnata ◽  
Length Variation ◽  
Mitochondrial Dna Control Region

The organization of the mitochondrial DNA (mtDNA) control region (CR) of the autumnal moth, Epirrita autumnata, is described. The E. autumnata CR presents a distinct type of lepidopteran CR with domains of non-repetitive and repetitive sequences. The CRs show considerable length variation owing to a variable number of short ~29-bp sequence blocks that are repeated between 6 and 14 times in tandem. The organization of such a tandem array is unique among the insect CRs examined so far. Furthermore, the E. autumnata CR, which may reach 1075 bp in length, is considerably longer than previously reported lepidopteran CRs, which reach 311–499 bp in length. Like other lepidopteran CRs, the E. autumnata CR contains two long homopolymer runs that may be involved in mtDNA replication and (or) transcription.Key words: Lepidoptera, Epirrita autumnata, mitochondrial DNA, control region, tandem array.

scholarly journals Variable Number Tandem Repeats in the Mitochondrial DNA of Lentinula edodes

Genes ◽  
2019 ◽  
Vol 10 (7) ◽  
pp. 542
Author(s):  
Kim ◽  
Song ◽  
Ha ◽  
Moon ◽  
Kim ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Dna Sequences ◽  
Tandem Repeats ◽  
Lentinula Edodes ◽  
Mechanistic Model ◽  
Variable Number ◽  
Type I ◽  
Complementary Sequence ◽  
Repeat Units ◽  
Variable Number Tandem Repeats

Variable number tandem repeats (VNTRs) in mitochondrial DNA (mtDNA) of Lentinula edodes are of interest for their role in mtDNA variation and their application as genetic marker. Sequence analysis of three L. edodes mtDNAs revealed the presence of VNTRs of two categories. Type I VNTRs consist of two types of repeat units in a symmetric distribution, whereas Type II VNTRs contain tandemly arrayed repeats of 7- or 17-bp DNA sequences. The number of repeat units was variable depending on the mtDNA of different strains. Using the variations in VNTRs as a mitochondrial marker and the A mating type as a nuclear type marker, we demonstrated that one of the two nuclei in the donor dikaryon preferentially enters into the monokaryotic cytoplasm to establish a new dikaryon which still retains the mitochondria of the monokaryon in the individual mating. Interestingly, we found 6 VNTRs with newly added repeat units from the 22 mates, indicating that elongation of VNTRs occurs during replication of mtDNA. This, together with comparative analysis of the repeating pattern, enables us to propose a mechanistic model that explains the elongation of Type I VNTRs through reciprocal incorporation of basic repeat units, 5’-TCCCTTTAGGG-3’ and its complementary sequence (5’-CCCTAAAGGGA-3’).

Structure and evolution of the mitochondrial control region of the pollen beetle Meligethes thalassophilus (Coleoptera: Nitidulidae)

Genome ◽  
2008 ◽  
Vol 51 (3) ◽  
pp. 196-207 ◽  
Author(s):  
Emiliano Mancini ◽  
Alessio De Biase ◽  
Paolo Mariottini ◽  
Alessandro Bellini ◽  
Paolo Audisio
Keyword(s):  
Control Region ◽  
Tandem Repeats ◽  
Population Level ◽  
Mitochondrial Control Region ◽  
Variable Number ◽  
12S Rrna ◽  
Rrna Gene ◽  
Length Variation ◽  
Pollen Beetle ◽  
Repeat Elements

The organization of the mitochondrial DNA (mtDNA) control region (CR) of the pollen beetle Meligethes thalassophilus is described. This mtDNA CR represents the longest sequenced for beetles so far, since the entire nucleotide sequence ranges from ~5000 to ~5500 bp. The CR of M. thalassophilus is organized in three distinct domains: a conserved domain near the tRNAIle gene, a variable domain flanking the 12S rRNA gene, and a relatively large central tandem array made up of a variable number of ~170 bp repeats that is responsible for the intraspecific length variation observed. Like other CRs found in insects, the M. thalassophilus CR contains two long homopolymeric runs that may be involved in mtDNA replication. Furthermore, conserved stem-and-loop structures in the repetitive domain were identified and their possible role in generating length variation is examined. Intraspecific comparison of the tandem repeat elements of M. thalassophilus suggests mechanisms of concerted evolution leading to homogenization of the repetitive region. The utility of such an array of tandem repeats as a genetic marker for assessing population-level variability and evolutionary relationships among populations is discussed. Finally, the technical difficulties found in isolating the mtDNA CR in beetles are remarked upon.

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