Origin and spread of human mitochondrial DNA haplogroup U7

Human mitochondrial DNA haplogroup U is among the initial maternal founders in Southwest Asia and Europe and one that best indicates matrilineal genetic continuity between late Pleistocene hunter-gatherer groups and present-day populations of Europe. While most haplogroup U subclades are older than 30 thousand years, the comparatively recent coalescence time of the extant variation of haplogroup U7 (~16–19 thousand years ago) suggests that its current distribution is the consequence of more recent dispersal events, despite its wide geographical range across Europe, the Near East and South Asia. Here we report 267 new U7 mitogenomes that – analysed alongside 100 published ones – enable us to discern at least two distinct temporal phases of dispersal, both of which most likely emanated from the Near East. The earlier one began prior to the Holocene (~11.5 thousand years ago) towards South Asia, while the later dispersal took place more recently towards Mediterranean Europe during the Neolithic (~8 thousand years ago). These findings imply that the carriers of haplogroup U7 spread to South Asia and Europe before the suggested Bronze Age expansion of Indo-European languages from the Pontic-Caspian Steppe region

A homozygous nonsense thyroid peroxidase mutation (R540X) consistently causes congenital hypothyroidism in two siblings born to a consanguineous family

WOS: 000367653300010PubMed ID: 26777044Objective: Congenital hypothyroidism (CH) is the most common neonatal endocrine disorder, and mutations in the thyroid peroxidase (TPO) gene have been reported to cause the disease. Our aim in this study was to determine the genetic basis of CH in two affected children coming from a consanguineous family. Methods: First, we investigated the potential genetic linkage of the family to any known CH locus using microsatellite markers and then screened for mutations in the linked gene by Sanger sequencing. By using next-generation sequencing, we also checked if any other mutation was present in the remaining 10 causative CH genes. Results: The family showed potential linkage to the TPO gene, and we detected a homozygous nonsense mutation (R540X) in both cases. The two patients had total iodide organification defect (TIOD). Both the microsatellite marker haplotypes and the mutation segregated with the disease status in the family, i.e. all healthy subjects were either heterozygous carriers or homozygous wild-type, confirming the pathogenic nature of the mutation. Neither was the mutation present in any of the 400 control chromosomes nor were there any other mutations in the remaining causative CH genes. Conclusion: This study proves the pathogenicity of R540X mutation and demonstrates the strong genotype/phenotype correlation associated with this mutation. It also highlights the power of working with familial cases in revealing the molecular basis of CH and in establishing accurate genotype/phenotype relationships associated with disease causing mutations

Recombinant plasmid-based quantitative Real-Time PCR analysis of salmonella enterica serotypes and its application to milk samples

WOS: 000371368200011PubMed ID: 26820062The aim of the current study was to develop, a new, rapid, sensitive and quantitative Salmonella detection method using a Real-Time PCR technique based on an inexpensive, easy to produce, convenient and standardized recombinant plasmid positive control. To achieve this, two recombinant plasmids were constructed as reference molecules by cloning the two most commonly used Salmonella-specific target gene regions, invA and ttrRSBC The more rapid detection enabled by the developed method (21 h) compared to the traditional culture method (90 h) allows the quantitative evaluation of Salmonella (quantification limits of 10(1) CFU/ml and 10(0) CFU/ml for the invA target and the ttrRSBC target, respectively), as illustrated using milk samples. Three advantages illustrated by the current study demonstrate the potential of the newly developed method to be used in routine analyses in the medical, veterinary, food and water/environmental sectors: I - The method provides fast analyses including the simultaneous detection and determination of correct pathogen counts; II - The method is applicable to challenging samples, such as milk; III - The method's positive controls (recombinant plasmids) are reproducible in large quantities without the need to construct new calibration curves.Middle East Technical University (METU) [BAP-03-14-2010-05]This study was supported by Middle East Technical University (METU) research fund: BAP-03-14-2010-05

Performance comparison of Next Generation sequencing platforms

Next Generation DNA Sequencing technologies offer ultra high sequencing throughput for very low prices. The increase in throughput and diminished costs open up new research areas. Moreover, number of clinicians utilizing DNA sequencing keeps growing. One of the main concern for researchers and clinicians who are adopting these platforms is their sequencing accuracy. We compared three of the most commonly used Next Generation Sequencing platforms; Ion Torrent from Life Technologies, GS FLX+ from Roche and HiSeq 2000 from Illumina

Topical single-dose vascular endothelial growth factor has no effect on soft tissue healing

Background: Vascular endothelial growth factor (VEGF) production in dermal wounds has been evaluated for evidence that it plays a probable role in wound healing. Events such as increased vascular permeability and concentration of infl ammatory cells on the site of injury, produced by VEGF, were linked to tissue repair. Aim: The present study aimed to evaluate the effects of single-dose topical administration of VEGF on wound healing. Materials and Methods: A total of 30 male Wistar albino rats weighing 250-280 g were used in this study. In addition, 2-cm-long skin incisions were created over bilaterally exposed skin of the tibia region in each rat. VEGF plasmid 2 µg was administered locally into the right side wound bed of each animal. No other procedure besides skin closure was administered on the left side. To determine histologic assessments, skin samples were obtained from six anesthetized rats at each interval (4, 8, 12, 16 and 30 days) through excisional biopsy. The tissues were fi xed in 10% neutral-buffered formalin for 1 week and then embedded in paraffi n wax. Transverse sections of the embedded tissue 5-7 µm thick were stained with hematoxylin and eosin (H and E). Results: There was no signifi cant difference regarding necrosis, epithelialization, infl ammation, fi broblast activity, ulcerative formation, or hemorrhage between experimental and control groups. No statistically signifi cant difference was found between the groups regarding granulation tissue formation and epidermal thickness. Conclusion: The administration method and dosage of VEGF is a major factor in terms of its effectiveness. The results of the present study did not evaluate the effectiveness of single-dose 2 µg topical administration of VEGF; however, various doses of VEGF plasmid should be tested in future studies in order to provide benefi cial effects from topical administration of VEGF

Determination of a new mutation in MT-ND1 gene of a patient with dextrocardia, ventriculoarterial discordance, and tricuspid atresia

Tricuspid atresia, a congenital heart defect (CHD) with unknown etiology, occurs 0.056 per 100 live births and is invariably associated with right ventricular hypoplasia, and atrial/ventricular septal defects (1). Recently, many of associations have been reported between mitochondrial DNA (mtDNA) and a variety of diseases (2). Next-generation sequencing has overcome many limitations of mtDNA studies, such as estimation of heteroplasmy level and its effects on the severity of mitochondrial diseases

Identification of red blood cell membrane defects in a patient with hereditary spherocytosis using next-generation sequencing technology and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

WOS: 000465868800060PubMed ID: 30896804Hereditary spherocytosis (HS) is characterized by the morphological transformation of erythrocytes into a spherical shape due to a hereditary defect in cell membrane proteins (ghosts) associated with disruption of erythrocyte skeletal structures. Contrary to the literature, pores were detected in the erythrocytes of a patient with HS. The aim of the present study was to determine the affected proteins and genes that were responsible for the pores. Ghost isolation was performed to determine the proteins responsible for the pores observed on the erythrocytes of the patient. Erythrocyte membrane proteins were visualized using SDS-PAGE. Exome and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS) analyses were used to identify the genes and proteins responsible for the observed defect. Quantitative protein assessments were performed using MALDI TOF MS. A difference was detected in the components of the erythrocyte membrane proteins. Band 3 and protein 4.2, which serve a particular role in membrane structure, decreased 4.573 and 4.106 fold, respectively. Through proteomic analyses, a non-synonymous exonic mutation region was identified in the Golgi membrane protein 1 (GOLM1) gene (Chr9 rs142242230). Sorting Intolerant From Tolerant and Polymorphism Phenotyping Scores, Likelihood Ratio Tests and MutationTaster revealed that the mutation was deleterious. The pores observed in the morphology of the erythrocytes may have developed due to the decrease in these proteins, which reside in the erythrocyte membrane structure. Furthermore, genetic profiling of the patient with HS and her family was conducted in the present study. Next-generation sequencing was used, and the genetic source of HS was identified as a GOLM1 gene mutation. The assessment of specific molecular defects is often not performed as the majority of mutations are unique to a family. However, molecular analyses should be performed in severe cases where prenatal diagnosis is required, or for unique HS phenotypes to aid scientific investigation.Istanbul University [35214]The present study was supported by the research fund of Istanbul University (grant no. 35214)