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SUMMARY: Here we introduce a Fiji plugin utilizing the HPC-as-a-Service concept, significantly mitigating the challenges life scientists face when delegating complex data-intensive processing workflows to HPC clusters. We demonstrate on a common Selective Plane Illumination Microscopy image processing task that execution of a Fiji workflow on a remote supercomputer leads to improved turnaround time despite the data transfer overhead. The plugin allows the end users to conveniently transfer image data to remote HPC resources, manage pipeline jobs and visualize processed results directly from the Fiji graphical user interface. AVAILABILITY AND IMPLEMENTATION: The code is distributed free and open source under the MIT license. Source code: https://github.com/fiji-hpc/hpc-workflow-manager/, documentation: https://imagej.net/SPIM_Workflow_Manager_For_HPC. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
BACKGROUND: A growing number of crystal and NMR structures reveals a considerable structural polymorphism of DNA architecture going well beyond the usual image of a double helical molecule. DNA is highly variable with dinucleotide steps exhibiting a substantial flexibility in a sequence-dependent manner. An analysis of the conformational space of the DNA backbone and the enhancement of our understanding of the conformational dependencies in DNA are therefore important for full comprehension of DNA structural polymorphism. RESULTS: A detailed classification of local DNA conformations based on the technique of Fourier averaging was published in our previous work. However, this procedure requires a considerable amount of manual work. To overcome this limitation we developed an automatic classification method consisting of the combination of supervised and unsupervised approaches. A proposed workflow is composed of k-NN method followed by a non-hierarchical single-pass clustering algorithm. We applied this workflow to analyze 816 X-ray and 664 NMR DNA structures released till February 2013. We identified and annotated six new conformers, and we assigned four of these conformers to two structurally important DNA families: guanine quadruplexes and Holliday (four-way) junctions. We also compared populations of the assigned conformers in the dataset of X-ray and NMR structures. CONCLUSIONS: In the present work we developed a machine learning workflow for the automatic classification of dinucleotide conformations. Dinucleotides with unassigned conformations can be either classified into one of already known 24 classes or they can be flagged as unclassifiable. The proposed machine learning workflow permits identification of new classes among so far unclassifiable data, and we identified and annotated six new conformations in the X-ray structures released since our previous analysis. The results illustrate the utility of machine learning approaches in the classification of local DNA conformations.
- MeSH
- algoritmy MeSH
- DNA chemie MeSH
- G-kvadruplexy MeSH
- klasifikace metody MeSH
- konformace nukleové kyseliny MeSH
- krystalografie rentgenová MeSH
- nukleární magnetická rezonance biomolekulární MeSH
- průběh práce MeSH
- shluková analýza MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVE: Restorative treatment of anterior teeth is often required as the final step of orthodontic therapy to optimize the esthetics and function. This case report presents a direct minimally invasive approach for post-orthodontic recontouring of anterior teeth using the composite injection technique with a digital workflow. CLINICAL CONSIDERATIONS: The extraction of central incisors with short roots was indicated to resolve maxillary anterior crowding. The gained space was closed with lateral incisors, and it was necessary to recontour them and canines to resemble central and lateral incisors, respectively. The restorations were digitally designed, and a transparent silicone index was fabricated from a 3D-printed cast of the digital wax-up. After lateral incisors and canines were bonded with a universal adhesive in the etch-and-rinse mode, a highly filled injectable composite resin was applied into the index. The restorative treatment was preceded by gingivectomy using an Er:YAG laser and home bleaching. CONCLUSIONS: In cases where more complex recontouring of anterior teeth is necessary, the composite injection technique could be a suitable alternative to indirect restorations, because it is straightforward, cost-effective, and does not require any preparation. The digital workflow simplified and expedited the treatment while contributing to its precision. CLINICAL SIGNIFICANCE: Post-orthodontic recontouring using direct composite restorations is well accepted by patients due to low cost and non-invasiveness. However, in complicated cases, the free-hand technique is time-consuming and technique-sensitive. The presented composite injection technique with a digital workflow significantly simplifies and expedites the composite placement while predictably enhancing the treatment outcome.
- MeSH
- estetika stomatologická MeSH
- lidé MeSH
- průběh práce * MeSH
- řezáky MeSH
- složené pryskyřice MeSH
- trvalá sanace zubů * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
Mycoplasma spp. contamination is a major concern in laboratories handling cell cultures, and routine detection methods are usually time-consuming, laborious and lack sensitivity. This study presents a streamlined workflow integrating rapid thermal DNA extraction (99 °C-1 min) with a SYBR Green-based qPCR for Mycoplasma detection. High-coverage primers targeting an 86-bp region of the 16S rDNA were designed using 109 Mycoplasma spp. sequences from GeneBank. In silico analysis confirmed full primer annealing to major cell culture contaminants (M. arginini, M. hominis, M. orale, and M. hyorhinis). Upon thermal lysis and qPCR optimization, the yield of the protocol was equivalent to that of phenol-chloroform extraction plus qPCR, with a detection limit of 64 bacterial cells. Finally, the performance of the protocol was confirmed in cell cultures with known Mycoplasma spp. contamination, accurately reproducing the contamination status. Thus, the developed protocol provides a simple, rapid, cost-effective, and sensitive method for monitoring Mycoplasma spp. in cell cultures.
- MeSH
- buněčné kultury * metody MeSH
- DNA bakterií genetika izolace a purifikace MeSH
- DNA primery genetika MeSH
- kvantitativní polymerázová řetězová reakce * metody MeSH
- lidé MeSH
- Mycoplasma * genetika izolace a purifikace klasifikace MeSH
- průběh práce MeSH
- ribozomální DNA genetika MeSH
- RNA ribozomální 16S genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Introduction: Circulating tumor DNA (ctDNA) has become a relevant biomarker in cancer management, allowing tumor assessment through analysis of minimally invasive liquid biopsies. Applications include screening, diagnostics, monitoring of treatment efficacy and detection of minimal residual disease as well as relapse. The potential of ctDNA analysis is significant, but several biological and technical challenges need to be addressed before widespread clinical implementation.Areas covered: Several clinical applications where ctDNA analysis may be beneficial require detection of individual DNA molecules. Consequently, to acquire accurate and informative data the entire workflow from sampling to final data interpretation needs to be optimized. In this review, we discuss the biological and technical challenges of ctDNA analysis and how preanalytical and analytical approaches affect different cancer applications.Expert opinion: While numerous studies have demonstrated the potential of using ctDNA in cancer applications, yet few reports about true clinical utility exist. Despite encouraging data, the sensitivity of ctDNA analyses, i.e. the probability to detect presence of cancer in liquid biopsies, is still an issue. Analysis of multiple mutations in combination with simultaneous assessment of other analytes is one solution. Improved standardization and guidelines will also facilitate the introduction of ctDNA analysis into clinical routine.
The limited availability of biological samples hinders phylogenetic efforts to define structural differences among various biological groups. A novel workflow enabling the analysis of protists in low cell numbers by electron microscopy (EM) is described with cysts of Giardia intestinalis, a single-celled eukaryotic parasite. Correlative light and electron microscopy (CLEM) allows for the selection of individual cells and is economical in terms of time and cost. We describe a cyst purification protocol in combination with an adhesive coating for fixation and ultrathin embedding that results in excellent preservation of cell morphology. The application of advanced structural and analytical EM methods, such as high-resolution field emission scanning electron microscopy (FESEM), focused ion beam tomography (FIB/SEM), and energy-dispersive X-ray spectroscopy (EDX) analysis, is demonstrated. The workflow represents a new approach for studying the cellular and organelle architecture of rare and "difficult to culture" microorganisms.
The enzymatic degradation of polyethylene terephthalate (PET) by PETases has gained significant attention as a potential solution for plastic waste management. However, the absence of a standardized protocol for PETase production across studies presents a challenge for consistent enzyme characterization and activity comparison. Variations in production methods, including expression systems and purification techniques, may contribute to discrepancies in reported PETase activities. Here, we present the development of a unified protocol for the production of wild-type and engineered IsPETase variants. This protocol comprises standardized expression, purification, and quality control steps to ensure reproducibility and reliability. By enabling more accurate comparisons of PETase variants and addressing inconsistencies in PETase production, this approach facilitates collaborative efforts to advance plastic degradation technologies and lays the groundwork for accelerating research in enzymatic PET degradation and its applications in plastic waste management.
