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Molecular and Cellular Probes

Qingyuan Huang, Chao Ye, Tingting Chen, Jiali Jiang, Yuanyi Peng, Jing Chen, Rendong Fang
Enzootic nasal tumor virus 2 (ENTV-2), the aetiological agent of enzootic nasal adenocarcinoma in goats, is prevalent in China; resulting in substantial economic losses to the goat-breeding industry. Therefore, it is necessary to establish an efficient detection method for the diagnosis and prevention of ENTV-2 infection. More recently, EvaGreen is emerging as a novel alternative fluorescent dye for quantitative real-time PCR because of its low cost, specific amplification and high resolution. In this study, we developed a specific, sensitive, and cost-effective detection method-an EvaGreen-based real-time PCR assay for the detection of ENTV-2...
February 13, 2019: Molecular and Cellular Probes
Hao-Ying Han, Hui-Hua Zheng, Yu Zhao, Run-Bo Tian, Peng-Li Xu, Hua-Lin Hou, Hong-Ying Chen, Ming-Fan Yang
The development of a rapid, specific, and sensitive SYBR Green I-based duplex real-time quantitative PCR assay is described for the simultaneous detection of porcine epidemic diarrhea virus (PEDV) and porcine circovirus type 3 (PCV3). The assay specifically detected PEDV and PCV3, with no fluorescence detected for other non-targeted pig pathogens. The assay showed a good linear relationship, and the limits of detection for this assay were 34.6 copies/μL and 61.2 copies/μL for PEDV and PCV3, respectively. The assay exhibited high repeatability and reproducibility, with intra-assay and inter-assay variation coefficients less than 2...
February 5, 2019: Molecular and Cellular Probes
Hui Liu, Zengfeng Ni, Lili Shi, Lijie Ma, Jianqiang Zhao
AIM: Studies have reported that micro (miR)-486-5p plays a crucial part in the progression of leukemia, however, to the best of our knowledge, few studies have been conducted on its mechanism in leukemia. In this study, the mechanism of miR-486-5p in leukemia cells was pointed out and its possible target genes were analyzed for the purpose of providing new therapeutic strategies for treating leukemia patients. METHODS: MiRNA expression of Leukemia cells (K562, Kasumi-1, and THP-1) and primary leukocytes was detected by Real-time Quantitative polymerase chain reaction(qPCR)...
February 4, 2019: Molecular and Cellular Probes
Yingying Zhang, Yaning Wang, Hongbao Wang, Xueyao Ma, Linsen Zan
In this study, the potential functions of miR-224 in regulating adipogenic differentiation were explored in bovine preadipocytes. Comparative transcriptome analysis between castrated male cattle with increased intramuscular fat (IMF) and intact male cattle revealed that miR-224 and LPL were abnormally expressed, correlating negatively, and LPL was a predicted target of miR-224. A dual luciferase reporter assay confirmed the negative targeting regulatory relationship between miR-224 and LPL. When miR-224 was either overexpressed or silenced, qRT-PCR showed a negative regulatory effect on LPL...
January 28, 2019: Molecular and Cellular Probes
Xiaoyan Yu, Yan Zhang, Ping Yang, Xueli Gao, Yuping Wang
Preeclampsia (PE), a special type of hypertensive disorder complicating pregnancy (HDCP), is highly associated with the migratory and invasive capacity of extravillous trophoblasts (EVTs). Here, we aimed to study the functions of low-density lipoprotein receptor-related protein 6 (LRP6) in PE pathogenesis. A comparative analysis of cellular gene expressions between placenta tissues collected from PE patients and normal pregnant women showed that the expressions of LRP6, β-catenin and matrix metallopeptidases/TIMP metallopeptidase inhibitors (MMPs/TIMPs) ratio in placentas of PE patients were much lower than the normal...
January 23, 2019: Molecular and Cellular Probes
Isabel Diebold, Ulrike Schön, Rita Horvath, Oliver Schwartz, Elke Holinski-Feder, Heike Kölbel, Angela Abicht
The heterooctameric mitochondrial trifunctional protein (MTP), composed of four α- and β-subunits harbours three enzymes that each perform a different function in mitochondrial fatty acid β-oxidation. Pathogenic variants in the MTP genes (HADHA and HADHB) cause MTP deficiency, a rare autosomal recessive metabolic disorder characterized by phenotypic heterogeneity ranging from severe, early-onset, cardiac disease to milder, later-onset, myopathy and neuropathy. Since metabolic myopathies and neuropathies are a group of rare genetic disorders and their associated muscle symptoms may be subtle, the diagnosis is often delayed...
January 22, 2019: Molecular and Cellular Probes
Yanting Lu, Xiuli Ma, Qinghua Kong, Yuhui Xu, Jing Hu, Fang Wang, Wanying Qin, Libin Wang, Wenyong Xiong
Insulin-responsive glucose transporter type 4 (GLUT4) translocation plays a major role in controlling glucose uptake in adipose tissue and muscle, maintaining homeostasis and preventing hyperglycemia. Screening for chemicals enhancing GLUT4 translocation is an approach for identifying hits of drug development for type 2 diabetes. Here we developed a novel functional dual-color probe, pHluorin-GLUT4-mOrange2, and constructed 3T3-L1 adipocytes based screening system to simply and efficiently screen new compounds stimulating GLUT4 translocation...
January 10, 2019: Molecular and Cellular Probes
Lea Neuheuser, Robert Meyer, Matthias Begemann, Miriam Elbracht, Thomas Eggermann
Imprinting Disorders are a group of rare diseases with overlapping phenotypes which are associated with similar molecular changes and affect imprinted chromosomal regions. Clinical features mainly occur prenatally or in childhood, but have a severe lifelong impact on health. Due to their clinical and molecular heterogeneity, the diagnosis of imprinting disorders is often challenging and requires testing of a broad spectrum of genomic variants and aberrant methylation of imprinted loci (epimutations). A significant number of patients suspicious for imprinting disorders remain without a molecular confirmation, and in these cases differential diagnoses have to be considered...
January 2, 2019: Molecular and Cellular Probes
Jianhu Pang, Qiong Wang, Yuejun Fei, Peng Zhu, Longliang Qiao, Hailong Huang, Chenyang Dang, Weifang Gao
Vibrio harveyi is a pathogen that infects fish and shellfish worldwide, causing severe economic losses for the aquaculture industry. As the early diagnosis of V. harveyi infection is crucial to disease surveillance and prevention in cultured marine animals, a fast and accurate method to detect V. harveyi is required. Here, we performed recombinase polymerase amplification (RPA) using novel primers specifically designed to recognize the V. harveyi toxR gene, which encodes a transmembrane protein, and then hybridized this gene with a carboxy fluorescein (FAM)-labeled probe...
January 2, 2019: Molecular and Cellular Probes
Mark Jain, Alexander Vladimirovich Balatsky, Daria Borisovna Revina, Larisa Mikhailovna Samokhodskaya
BACKGROUND: Blood of pregnant women contains cell-free fetal DNA (cffDNA), which is widely used in non-invasive prenatal diagnosis. The modern laboratory equipment market provides huge variety of commercial kits for isolation of circulating nucleic acids, but unfortunately none of them are standardized for isolation of cffDNA, which is a crucial step for success of subsequent analysis. AIM: To compare DSPVK and CNAK in terms of cffDNA, cell-free total DNA (cftDNA) yield and resulting cffDNA fraction, as well as to try to explain the possible difference between the efficacy of these kits...
December 22, 2018: Molecular and Cellular Probes
Rong Zeng, Jinyan Luo, Shigang Gao, Lihui Xu, Zhiwei Song, Fuming Dai
Reverse transcription recombinase polymerase amplification (RT-RPA) for detection of Cucumber green mottle mosaic virus (CGMMV) was developed in this study. It was found to be specific, with a limit of detection of 0.5 pg of total CGMMV RNA. CGMMV on inoculated leaves was tested using RT-RPA assay, suggesting that this method is suitable for CGMMV detection from leaves.
December 21, 2018: Molecular and Cellular Probes
Roberto Cruz-Flores, Hung Nam Mai, Arun K Dhar
Acute hepatopancreatic necrosis disease (AHPND), also known as Early mortality syndrome (EMS), is a recently emerged lethal disease that has caused major economic losses in shrimp aquaculture. The etiologic agents are Vibrio spp. that carry Photorhabdus Insect-Related (Pir) toxin genes pirA and pirB. A multiplex SYBR Green real-time PCR was developed that detects pirA, pirB, and two internal control genes, the shrimp 18S rRNA and the bacterial 16S rRNA genes in a single reaction. The pirB primers amplify the 3'-end of the pirB gene allowing the detection of Vibrio spp...
December 18, 2018: Molecular and Cellular Probes
Han Zhao, Changbei Ma, Mingjian Chen
In this paper, we propose a fluorometric approach for the highly sensitive detection of inorganic pyrophosphatase (PPase) based on G-quadruplex-thioflavin T (ThT). In the absence of PPase, Cu2+ can coordinate with pyrophosphate (PPi) to generate a Cu2+ /PPi complex. Then the G-rich sequence folds into the G-quadruplex structure, which can combine with ThT to generate a remarkable fluorescent signal. In the presence of PPase, the coordinated compound can be destroyed by the PPase catalyzed hydrolysis of PPi into inorganic phosphate (Pi)...
December 17, 2018: Molecular and Cellular Probes
Atcharaporn Kidchana, Nattakan Meekhanon, Rujirat Hatrongjit, Marcelo Gottschalk, Anusak Kerdsin
Random amplification of polymorphic DNA (RAPD) and 16S-23S rDNA intergenic spacer polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were applied and evaluated to determine clonal complexes (CCs) of 684 Streptococcus suis isolates from pigs and humans. RAPD better distinguished major S. suis CCs than the PCR-RFLP method. The assay was capable of simultaneously distinguishing CC1, CC16, CC25, CC28, CC104, CC221/234, and CC233/379. PCR-RFLP could not clearly differentiate among most CCs in this study except CC16...
December 5, 2018: Molecular and Cellular Probes
Ping Zhou, Weijie Xie, Yifan Sun, Ziru Dai, Guang Li, Guibo Sun, Xiaobo Sun
Mitochondria play a central role in various critical cellular processes, including energy synthesis, energy supply and apoptosis. Panax notoginseng, a commonly used traditional Chinese medicine, has various pharmacological effects on the human body. Ginsenosides are representative bioactive components of P. notoginseng. Recently, more attention has focused on ginsenoside Rb1 as an antioxidative and anti-inflammatory agent that can protect the nervous system and the cardiovascular system. Numerous studies have shown that Rb1 exerts these effects by regulating mitochondrial energy metabolism, mitochondrial fission and fusion, apoptosis, oxidative stress and reactive oxygen species release, mitophagy and mitochondrial membrane potential...
December 4, 2018: Molecular and Cellular Probes
Malik Siddique Mahmood, Muhammad Asad Ullah, Hina Batool, Sana Batool, Naeem Mehmood Ashraf
Gonorrheal infections are the second most prevalent sexually transmitted diseases STDs in the USA populations after Chlamydia. These infections have now become an urgent problem to address because Neisseria gonorrhoeae is capable of developing resistance to multiple antibiotic classes. Infection with these antibiotic-resistant strains has become the major public health concern. Although extensive researches are ongoing to control its transmission and to develop the productive treatments against this pathogen, no effective vaccine could develop till now...
November 28, 2018: Molecular and Cellular Probes
Sadaomi Sugimoto, Hiroshi Maeda, Mizuki Kitamatsu, Ikuo Nishikawa, Muneyasu Shida
Peptide nucleic acids (PNA) are DNA/RNA analogs in which the sugar-phosphate backbone is replaced by N-2-aminoethylglycine. PNA are widely used for experimental antisense therapy due to their strong affinity to mRNA. By targeting specific genes, protein synthesis and the growth of bacteria or cancer cells can be inhibited by PNA. Here, we report the design and evaluation of antisense PNA for selective growth inhibition of Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, potent pathogens associated with periodontitis...
November 21, 2018: Molecular and Cellular Probes
Yongning Zhang, Zhou Zhang, Zhanying Wang, Zili Wang, Caixia Wang, Chunyan Feng, Wanzhe Yuan, Xiangmei Lin, Shaoqiang Wu
Porcine circovirus 3 (PCV3), a newly emerged circovirus, is associated with porcine dermatitis and nephropathy syndrome, reproductive failure and multi-systemic inflammation disease, and is widely distributed in pig populations worldwide. Therefore, developing specific diagnostic assays will be important for controlling this emerging pathogen. In this study, we developed a novel droplet digital PCR (ddPCR) assay targeting the PCV3 cap gene to improve the sensitivity of PCV3 detection. The established assay is highly specific to PCV3, and does not cross react with other important swine pathogens...
November 20, 2018: Molecular and Cellular Probes
Claudia Techera, Gonzalo Tomás, Yanina Panzera, Alejandro Banda, Paula Perbolianachis, Ruben Pérez, Ana Marandino
Infectious bursal disease virus (IBDV) and chicken anemia virus (CAV) cause relevant immunosuppressive diseases in poultry. Clinical diagnosis of these viruses is challenging given the different disease presentations and the frequent occurrence of co-infections with other pathogens. Here, we standardized and validated simplex and duplex RT-qPCR assays for the straightforward detection of IBDV and CAV. The qPCR assays are based on primers and hydrolysis probes that target highly conserved regions of IBDV and CAV genomes...
November 14, 2018: Molecular and Cellular Probes
Shuyi Han, Huili Yang, Yunhui Han, Hongzhi Zhang
PURPOSE: Maternal type I diabetes mellitus (T1DM) increases the risk of adverse pregnancy outcomes, but the corresponding mechanism is unclear. This study aims to investigate the mechanism underlying the adverse pregnancy outcomes of maternal T1DM. METHODS: Gene expression microarray (GSE51546) was down-loaded from the Gene Expression Omnibus. This dataset included 12 umbilical cord samples from the newborns of T1DM mothers (T1DM group, N = six) and non-diabetic mothers (control group, N = six)...
November 14, 2018: Molecular and Cellular Probes
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