T Oligo-Primed Polymerase Chain Reaction (TOP-PCR): A Robust Method for the Amplification of Minute DNA Fragments in Body Fluids.

Body fluid DNA sequencing is a powerful noninvasive approach for the diagnosis of genetic defects, infectious agents and diseases. The success relies on the quantity and quality of the DNA samples. However, numerous clinical samples are either at low quantity or of poor quality due to various reasons. To overcome these problems, we have developed T oligo-primed polymerase chain reaction (TOP-PCR) for full-length nonselective amplification of minute quantity of DNA fragments. TOP-PCR adopts homogeneous "half adaptor" (HA), generated by annealing P oligo (carrying a phosphate group at the 5' end) and T oligo (carrying a T-tail at the 3' end), for efficient ligation to target DNA and subsequent PCR amplification primed by the T oligo alone. Using DNA samples from body fluids, we demonstrate that TOP-PCR recovers minute DNA fragments and maintains the DNA size profile, while enhancing the major molecular populations. Our results also showed that TOP-PCR is a superior method for detecting apoptosis and outperforms the method adopted by Illumina for DNA amplification

Determination of Nucleopolyhedrovirus’ Taxonomic Position

To date , over 78 genomes of nucleopolyhedroviruses (NPVs) have been sequenced and deposited in NCBI. How to define a new virus from the infected larvae in the field is usually the first question. Two NPV strains, which were isolated from casuarina moth (L. xylina) and golden birdwing larvae (Troides aeacus), respectively, displayed the same question. Due to the identity of polyhedrin (polh) sequences of these two isolates to that of Lymantria dispar MNPV and Bombyx mori NPV, they are named LdMNPV-like virus and TraeNPV, provisionally. To further clarify the relationships of LdMNPV-like virus and TraeNPV to closely related NPVs, Kimura 2-parameter (K-2-P) analysis was performed. Apparently, the results of K-2-P analysis that showed LdMNPV-like virus is an LdMNPV isolate, while TraeNPV had an ambiguous relationship to BmNPV. Otherwise, MaviNPV, which is a mini-AcMNPV, also exhibited a different story by K-2-P analysis. Since K-2-P analysis could not cover all species determination issues, therefore, TraeNPV needs to be sequenced for defining its taxonomic position. For this purpose, different genomic sequencing technologies and bioinformatic analysis approaches will be discussed. We anticipated that these applications will help to exam nucleotide information of unknown species and give an insight and facilitate to this issue

Significant Inhibition of Tumor Growth following Single Dose Nanoparticle-Enhanced Photodynamic Therapy

Photodynamic therapy (PDT) for cancer treatment involves the pathology’s uptake of photosensitizers, which produce cytotoxic reactive oxygen species by photoirradiation. The use of nanoparticles as carriers of photosensitizers is one promising approach to this endeavor, owing to their small size, unique physicochemical properties, and easy/diverse functionalization. In the current work, we report on the in vivo assessment of PDT efficacy of these nanoconstructs in a murine model of human breast cancer, following a single (one-shot) nanoparticle dose and photoirradiation. Palladium-porphyrin (PdTPP) was administered intratumorally via injection of aqueous suspensions of either free PdTPP or MSN-conjugated PdTPP (MSN-PdTPP) at a dose of 50 μg. Mice were then exposed to a single photoirradiation session with total energy of 80 J. One month after one-shot PDT treatment, significantly greater reductions in tumor growth were observed in MSN-Pd treated animals than in PdTPP cohorts. Electron microscopy of tumor specimens harvested at various timepoints revealed excellent MSN-PdTPP uptake by cancer cells while immunohistologic analysis demonstrated marked increases in apoptotic response of MSN-PdTPP treated animals relative to PdTPP controls. Taken together, these findings suggest that considerable improvements in PDT efficacy can readily be achieved via the use of nanoparticle-based photosensitizers

Production of Active Nonglycosylated Recombinant B-Chain of Type-2 Ribosome-Inactivating Protein from Viscum articulatum and Its Biological Effects on Peripheral Blood Mononuclear Cells

Type-2 ribosome-inactivating proteins, composed of a toxic A-chain and lectin-like B-chain, display various biological functions, including cytotoxicity and immunomodulation. We here cloned the lectin-like B-chain encoding fragment of a newly identified type-2 RIP gene, articulatin gene, from Viscum articulatum, into a bacterial expression vector to obtain nonglycosylated recombinant protein expressed in inclusion bodies. After purification and protein refolding, soluble refolded recombinant articulatin B-chain (rATB) showed lectin activity specific toward galactoside moiety and was stably maintained while stored in low ionic strength solution. Despite lacking glycosylation, rATB actively bound leukocytes with preferential binding to monocytes and in vitro stimulated PBMCs to release cytokines without obvious cytotoxicity. These results implicated such a B-chain fragment as a potential immunomodulator

Image operator learning coupled with CNN classification and its application to staff line removal

Many image transformations can be modeled by image operators that are characterized by pixel-wise local functions defined on a finite support window. In image operator learning, these functions are estimated from training data using machine learning techniques. Input size is usually a critical issue when using learning algorithms, and it limits the size of practicable windows. We propose the use of convolutional neural networks (CNNs) to overcome this limitation. The problem of removing staff-lines in music score images is chosen to evaluate the effects of window and convolutional mask sizes on the learned image operator performance. Results show that the CNN based solution outperforms previous ones obtained using conventional learning algorithms or heuristic algorithms, indicating the potential of CNNs as base classifiers in image operator learning. The implementations will be made available on the TRIOSlib project site.Comment: To appear in ICDAR 201

Dual Targeting of 3-Hydroxy-3-methylglutaryl Coenzyme A Reductase and Histone Deacetylase as a Therapy for Colorectal Cancer

AbstractStatins are 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (HMGR) inhibitors decreasing serum cholesterol and have shown promise in cancer prevention. In this study, we demonstrated the oncogenic role of HMGR in colorectal cancer (CRC) by disclosing increased HMGR activity in CRC patients and its enhancement of anti-apoptosis and stemness. Our previous studies showed that statins containing carboxylic acid chains possessed activity against histone deacetylases (HDACs), and strengthened their anti-HDAC activity through designing HMGR-HDAC dual inhibitors, JMF compounds. These compounds exerted anti-cancer effect in CRC cells as well as in AOM-DSS and ApcMin/+ CRC mouse models. JMF mostly regulated the genes related to apoptosis and inflammation through genome-wide ChIP-on-chip analysis, and Ingenuity Pathways Analysis (IPA) predicted their respective regulation by NR3C1 and NF-κB. Furthermore, JMF inhibited metastasis, angiogenesis and cancer stemness, and potentiated the effect of oxaliplatin in CRC mouse models. Dual HMGR-HDAC inhibitor could be a potential treatment for CRC

Whole-genome DNA methylome analysis of different developmental stages of the entomopathogenic fungus Beauveria bassiana NCHU-157 by nanopore sequencing

The entomopathogenic fungus (EPF), Beauveria bassiana, is an important and commonly used EPF for microbial control. However, the role of DNA methylation has not been thoroughly studied. Therefore, the whole genomic DNA methylome of one promising EPF isolate, B. bassiana NCHU-157 (Bb-NCHU-157), was investigated by Oxford Nanopore Technologies (ONT). First, the whole genome of Bb-NCHU-157 was sequenced by next-generation sequencing (NGS) and ONT. The genome of Bb-NCHU-157 contains 16 contigs with 34.19 Mb and 50% GC content, which are composed of 10,848 putative protein-coding genes. Two putative DNA methyltransferases (DNMTs) were found, including Dim-2 and C-5 cytosine-specific DNA methylases. Both DNMTs showed higher expression levels in the mycelium stage than in the conidia stage, indicating that development of DNA methylation in Bb-NCHU-157 might occur in the mycelium stage. The global methylation level of the mycelium stage (5 mC = 4.56%, CG = 3.33%, CHG = 0.74%, CHH = 0.49%) was higher than that of the conidial stage (5 mC = 2.99%, CG = 1.99%, CHG = 0.63%, CHH = 0.37%) in both the gene and transposable element (TE) regions. Furthermore, the TE regions showed higher methylation frequencies than the gene regions, especially for CHH site methylation, suggesting regulation of genomic stabilization during mycelium development. In the gene regions, high methylation frequencies were found around the transcription start site (TSS) and transcription end site (TES). Moreover, CG and CHG methylation mainly occur in the promoter and intergenic regions, while CHH methylation occurs in the TE region. Among the methylated regions, 371, 661, and 756 differentially DNA methylated regions (DMRs) were hypermethylated in the mycelium in CG, CHG, and CHH, while only 13 and 7 DMRs were hypomethylated in the mycelium in CHG, and CHH, respectively. Genes located in the DMR shared the GO terms, DNA binding (GO: 0003677), and sequence-specific DNA binding (GO: 0043565) for hypermethylation in the mycelium, suggesting that methylation might regulate gene expression from the initial process. Evaluation of the DNA methylome in Bb-NCHU-157 by ONT provided new insight into this field. These data will be further validated, and epigenetic regulation during the development of B. bassiana will be explored