Mitochondrial DNA screening by melting curve analysis using peptide nucleic acid probes.

Analysis of single nucleotide polymorphisms (SNPs) in mitochondrial (mt)DNA hypervariable regions (HV) 1/2 is valuable in forensic investigations. We developed a method for mtDNA screening of the HV1 and HV2 regions by melting curve analysis, using peptide nucleic acid (PNA) probes. This method focuses on melting peak patterns obtained by thermal dissociation of PNA/DNA duplexes in amplified mtDNA products. Five PNA probe sets were designed to detect 25 SNPs in the two HV regions. We also detected non-target SNPs based on unexpected melting temperature (Tm) shifts. In fact, 62 SNPs (42 SNPs in HV1 and 20 in HV2) were identified, including the 25 target SNPs. Using this method, 46 melting peak patterns, including 8 pattern groups, were obtained in 60 unrelated individuals. The peak patterns were compared to 55 haplotypes identified by Sanger sequencing. The results obtained from analysis of target mtDNA SNPs were entirely consistent with those obtained by Sanger sequencing. Screening the HV1 and HV2 regions of mtDNA by this method may help minimize unnecessary recourse to full sequence analysis, allows to rapidly exclude samples that do not match evidence and reference samples, and may reduce turnaround times and analysis costs. Overall, this method may be effective and helpful in forensic investigations.

Analysis of ovarian volume of Korean children and adolescents at magnetic resonance imaging.

BACKGROUND: Knowledge of ovarian volume is important for diagnostic evaluations; however, normal ovarian volume studies on children and adolescents are lacking. OBJECTIVE: This study aimed to analyze age-specific ovarian volume and identify the diverse factors that contribute to ovarian diagnoses. MATERIALS AND METHODS: We retrospectively enrolled 180 patients (0-18 years of age) with normal ovaries who underwent magnetic resonance imaging (MRI) between 2010 and 2018. MRI sequences included coronal and axial T2-weighted turbo spin echo (TSE) images and coronal T1-weighted TSE images. Ovarian volume was calculated by the standard ellipsoid formula. Age-specific ovarian volume, height, weight, height-adjusted total ovarian volume and body mass index were obtained. Linear regression analysis was used to predict ovarian volume. RESULTS: Six age groups (infant; early and late child, and early, middle and late adolescent) were described. The early adolescent group (10-12 years) had the highest rate of increase. In the middle adolescent period (13-15 years), the curve of ovarian volume appeared flat. CONCLUSION: Our findings provide age-specific references for ovarian volume.

Characterization of Edge Contact: Atomically Resolved Semiconductor-Metal Lateral Boundary in MoS2.

Despite recent efforts for the development of transition-metal-dichalcogenide-based high-performance thin-film transistors, device performance has not improved much, mainly because of the high contact resistance at the interface between the 2D semiconductor and the metal electrode. Edge contact has been proposed for the fabrication of a high-quality electrical contact; however, the complete electronic properties for the contact resistance have not been elucidated in detail. Using the scanning tunneling microscopy/spectroscopy and scanning transmission electron microscopy techniques, the edge contact, as well as the lateral boundary between the 2D semiconducting layer and the metalized interfacial layer, are investigated, and their electronic properties and the energy band profile across the boundary are shown. The results demonstrate a possible mechanism for the formation of an ohmic contact in homojunctions of the transition-metal dichalcogenides semiconductor-metal layers and suggest a new device scheme utilizing the low-resistance edge contact.

A new method for ABO genotyping using fluorescence melting curve analysis based on peptide nucleic acid probes.

ABO genotyping has been routinely used to identify suspects or unknown remains in crime investigations. Probe-based fluorescence melting curve analysis (FMCA) is a powerful tool for mutation detection and is based on melting temperature shifts due to thermal denaturation. In the present study, we developed a new method for ABO genotyping using peptide nucleic acid (PNA) probe-based FMCA. This method allowed for the simultaneous detection of three single nucleotide polymorphism (SNP) sites in the ABO gene (nucleotide positions 261, 526, and 803) and the determination of 14 ABO genotypes (A/A, A/O01 or A/O02, A/O03, B/B, B/O01 or B/O02, B/O03, O01/O01 or O01/O02 or O02/O02, O01/O03 or O02/O03, O03/O03, A/B, cis-AB01/A, cis-AB01/B, cis-AB01/O01 or cis-AB01/O02, and cis-AB01/cis-AB01). Using this method, we analyzed 80 samples and successfully identified ABO genotypes (A/A [n=5], A/O01 or A/O02 [n=23], B/B [n=3], B/O01 or B/O02 [n=18], A/B [n=9], O01/O01 or O01/O02 or O02/O02 [n=20], cis-AB01/A [n=1], and cis-AB01/O01 or cis-AB01/O02 [n=1]). In addition, all steps in the method, including polymerase chain reaction, PNA probe hybridization, and FMCA, could be performed in one single closed tube in less than 3h. Since no processing or separation steps were required during analysis, this method was more convenient and rapid than traditional methods and reduced the risk of contamination. Thus, this method may be an effective and helpful tool in forensic investigations.

Source localization approach for functional DOT using MUSIC and FDR control.

In this paper, we formulate diffuse optical tomography (DOT) problems as a source localization problem and propose a MUltiple SIgnal Classification (MUSIC) algorithm for functional brain imaging application. By providing MUSIC spectra for major chromophores such as oxy-hemoglobin (HbO) and deoxy-hemoglobin (HbR), we are able to investigate the spatial distribution of brain activities. Moreover, the false discovery rate (FDR) algorithm can be applied to control the family-wise error in the MUSIC spectra. The minimum distance between the center of mass in DOT and the Montreal Neurological Institute (MNI) coordinates of target regions in experiments was between approximately 6 and 18 mm, and the displacement of the center of mass in DOT and fMRI ranged between 12 and 28 mm, which demonstrate the legitimacy of the DOT-based imaging. The proposed brain mapping method revealed its potential as an alternative algorithm to monitor the brain activation.

Lipid metabolic effect of Korean red ginseng extract in mice fed on a high-fat diet.

BACKGROUND: Ginseng saponin and ginsenosides exert anti-obesity effects via the modulation of physiological lipid metabolism in vivo or intracellular signalling in cell culture systems. However, the complicated relationship between the anti-obesity effects of ginseng and gene expression has yet to be defined under in vivo conditions. Therefore, we evaluated the relationship between the anti-obesity effects of Korean red ginseng extract (KRGE) and hepatic gene expression profiles in mice fed long-term on a high-fat diet (HFD) in this study. RESULTS: KRGE reduces the levels of cholesterol, low-density lipoprotein-cholesterol (LDL-C), serum triglycerides, and atherogenic indices. Levels of leptin, adiponectin and insulin, which regulate glucose and lipid metabolism, were impaired profoundly by HFD. However, KRGE treatment brought these levels back to normal. KRGE was found to down-regulate genes associated with lipid metabolism or cholesterol metabolism (Lipa, Cyp7a1, Il1rn, Acot2, Mogat1, Osbpl3, Asah3l, Insig1, Anxa2, Vldlr, Hmgcs1, Sytl4, Plscr4, Pla2g4e, Slc27a3, Enpp6), all of which were up-regulated by HFD. CONCLUSION: KRGE regulated the expression of genes associated with abnormal physiology via HFD. Leptin, insulin, and adiponectin, which carry out critical functions in energy and lipid metabolism, were shown to be modulated by KRGE. These results show that KRGE is effective in preventing obesity.

Anomalous origin of the left circumflex coronary artery from the first diagonal branch presented as acute myocardial infarction.

Coronary artery anomalies are diagnosed in 0.6 to 1.5% of patients who undergo coronary angiography (CAG). They may present with life threatening conditions but are generally asymptomatic. Recognition and adequate visualization of the anomaly is essential for correct management of the condition. However, in some cases the exact orifice and course of an anomalous coronary vessel cannot be selectively identified by CAG. In this report, a 54-year-old man was admitted to the hospital with acute inferior myocardial infarction and had an anomalous origin of the left circumflex coronary artery (LCX) from the first diagonal branch (D1). In CAG, the right CAG showed no significant stenosis and fortunately we found an anomalous origin of the LCX from the D1. The course of LCX was precisely established by 64-slice multi-detector computed tomography.

Development of a low-density DNA microarray for diagnosis of target-site mutations of pyrethroid and organophosphate resistance mutations in the whitefly Bemisia tabaci.

BACKGROUND: Rapid and accurate detection of mutations related to insecticide resistance is essential for development of resistance management strategies to support sustainable agriculture. The M918V, L925I and T929V mutations of the voltage-gated sodium channel gene (vgsc) and the F392W mutation of the acetylcholinesterase I gene (ace1) are reportedly associated with resistance to pyrethroids and organophosphates, respectively, in Bemisia tabaci. In order to detect known base substitutions in the ace1 and vgsc genes, a low-density microarray with an allele-specific probe was developed. RESULTS: Specific regions of the ace1 and vgsc gene mutations were amplified by multiplex asymmetrical PCR using Cy3-labelled primers, and then the PCR products were hybridised on the microarray. After analysing the probe signal data, the microarray containing 12 allele-specific probes produced a unique pattern of probe signals for field DNA samples of B. tabaci. To determine the optimal cut-off value of each probe, receiver operating characteristic (ROC) curve analysis was conducted using SPSS. Among 60 individual samples, microarray data for 57 samples were consistent with direct sequencing data. CONCLUSION: Although many molecular detection methods have been employed to monitor insecticide resistance, the present microarray provides rapid and accurate identification of target mutations in B. tabaci for resistance management.

A DNA microarray for identification of selected Korean birds based on mitochondrial cytochrome c oxidase I gene sequences.

DNA barcoding with the gene encoding cytochrome c oxidase I (COI) in the mitochondrial genome has been proposed as a standard marker to identify and discover animal species. Some migratory wild birds are suspected of transmitting avian influenza and pose a threat to aircraft safety because of bird strikes. We have previously reported the COI gene sequences of 92 Korean bird species. In the present study, we developed a DNA microarray to identify 17 selected bird species on the basis of nucleotide diversity. We designed and synthesized 19 specific oligonucleotide probes; these probes were arrayed on a silylated glass slide. The length of the probes was 19-24 bps. The COI sequences amplified from the tissues of the selected birds were labeled with a fluorescent probe for microarray hybridization, and unique hybridization patterns were detected for each selected species. These patterns may be considered diagnostic patterns for species identification. This microarray system will provide a sensitive and a high-throughput method for identification of Korean birds.

Usefulness of 64-slice multidetector computed tomography as an initial diagnostic approach in patients with acute chest pain.

BACKGROUND: Recently, multidetector computed tomography (MDCT) has been proposed as an accurate diagnostic tool to evaluate for coronary artery disease. However, the role of MDCT as part of the initial diagnostic for evaluating acute chest pain is less well established. METHODS: We prospectively enrolled patients presenting with acute chest pain to the emergency department (ED) and risk stratified them based on the pretest probability for an acute coronary syndrome (ACS): (1) very low, (2) low, (3) intermediate, (4) high, and (5) very high or definite. After exclusion of very low and very high risk patients, 268 patients were randomized to either immediate 64-slice cardiac MDCT or a conventional diagnostic strategy. Number of admissions, ED and hospital length of stay (LOS), and major adverse cardiac events over 30 days of follow-up were compared between the strategies based on the pretest probability for ACS. RESULTS: The number of patients ultimately diagnosed with an ACS did not differ between the 2 strategies. Emergency department LOS and total admissions were not different between strategies. Patients in the MDCT-based strategy had a decreased hospital LOS (P = .049) and fewer admissions deemed unnecessary (P = .007). Reductions in unnecessary admissions were more prominent in intermediate-risk patients (P = .015). None of the patients discharged from the ED in the MDCT-based strategy experienced major adverse cardiac events at follow-up. CONCLUSION: Use of an MDCT-based strategy in the ED as part of the initial diagnostic approach for patients presenting with acute chest pain is safe and efficiently reduces avoidable admissions in patients with an intermediate pretest probability for ACS.

Application of multiplex bead array assay for Yq microdeletion analysis in infertile males.

The purpose of this study was to apply the multiplex bead array as a diagnostic tool for male infertility. The multiplex bead array offers a new platform in high-throughput nucleic acid detection. Six loci, including sex-determining regions on the Y (SRY) chromosome as a control and five sequence-tagged sites (STS) in azoospermia-factor regions, were used in this system. Extracted genomic DNA from infertile male blood was used for multiplex polymerase chain reaction (PCR). After multiplex PCR using specific Cy3-labeled primer sets, the PCR product was hybridized with capture probes. A multiplexed PCR-liquid bead was arrayed for simultaneous detection using the Luminex system. This assay system correctly identified the presence or deletion of the Y chromosome. Therefore, this method provides a sensitive and high-throughput method for probing the deletion of the Y chromosome, and offers a completely new approach to male infertility screening.

Gene expression analysis of peroxisome proliferators- and phenytoin-induced hepatotoxicity using cDNA microarray.

The recent DNA microarray technology enables us to understand a large number of gene expression profiling. The technology has potential possibility to comprehend mechanism of multiple genes were related to compounds which have toxicity in biological system. So, the toxicogenomics through this technology may be very powerful for understanding the effect of unknown toxic mechanisms in biological system. We have studied that the effect of compounds related to hepatotoxin in vivo system using DNA microarray and classified chemicals which have been well characterized. We have studied three compounds; 2 peroxisome proliferators: Clofibrate (ethyl-p-chlorophenoxyisobutyrate), gemfibrozil (5-2[2,5-dimethyl-phenoxy]2-2-dimethyl-pentanonic), and an antiepileptic drug: phenytoin (5,5-diphenylhydantoin). Male Sprague-Dawely VAF(+) albino rats of 5-6 weeks old were treated with each compound for 24 hr and 2 weeks. 4.8 K cDNA microarray in house has been used for gene expression profiling. We found that the clustering of gene expression had similarity like as the toxic phenotype of compounds.