Cleavage of delta and epsilon tubulin PCR products is observed upon PCR purification.

In this study, we report an unusual phenomenon of the self-cleavage of purified PCR products of codon-optimized delta tubulin ( ) and epsilon tubulin ( ) genes through an unknown mechanism. Our studies revealed that intact PCR products for both these genes could be obtained upon PCR amplification from plasmid templates carrying these genes. However, interestingly, purification of these PCR products led to their cleavage through an unidentified mechanism.

Light intensity and spectral quality modulation for improved growth kinetics and biochemical composition of Chlamydomonas reinhardtii.

Effective strategies to optimize algal growth and lipid productivity are critical for the sustainable production of biomass for various applications. Light management has emerged as a promising approach, but the intricate relationship between light intensity, spectral quality, and algal responses remains poorly understood. This study investigated the effects of different light qualities (blue, red-orange, and white-yellow) and intensities (45-305 μmol/m·s) on Chlamydomonas reinhardtii.

Evaluation of AI Model for Cephalometric Landmark Classification (TG Dental).

The accuracy of cephalometric landmark identification for malocclusion classification is essential for diagnosis and treatment planning. Identifying these landmarks is often complex and time-consuming for orthodontists. An AI model for classification was recently developed.

LED alternating between blue and red-orange light improved the biomass and lipid productivity of Chlamydomonas reinhardtii.

Light management is important for improving algae cultivation, specifically by enhancing the productivity of biomass and valued bioproducts. In this study, we present evidence that alternating blue and red-orange light can improve the algal growth kinetics and lipid production in a photobioreactor. Blue (430-445, 460-470 nm) and red-orange light (580-660 nm) from a LED were set at the light saturation point (B: 65 μmol/ms; RO: 155 μmol/ms) and alternated for the cultivation of the green alga Chlamydomonas reinhardtii.

Interactive effects of light quality and culturing temperature on algal cell size, biomass doubling time, protein content, and carbohydrate content.

Light management strategy can be used to improve algal biomass and nutrient production. However, the response of algal metabolism to different light qualities, especially their interaction with other environmental factors, is not well understood. This study focuses on the interactive effects of light quality and culturing temperature on algal protein content and carbohydrate content of C.

Risks and Benefits of Glioblastoma Resection in Older Adults: A Retrospective Austrian Multicenter Study.

Objective: To assess the prognostic profile, clinical outcome, treatment-associated morbidity, and treatment burden of elderly patients with glioblastoma (GBM) undergoing microsurgical tumor resection as part of contemporary treatment algorithms.

Methods: We retrospectively identified patients with GBM ≥65 years of age who were treated by resection at 2 neuro-oncology centers. Survival was assessed by Kaplan-Meier analyses; log-rank tests identified prognostic factors.

Evaluation of endoscopic visible light spectroscopy: comparison with microvascular oxygen tension measurements in a porcine model.

Background: Visible light spectroscopy (VLS) is a technique used to measure the mucosal oxygen saturation during upper gastrointestinal endoscopy to evaluate mucosal ischemia, however in vivo validation is lacking. We aimed to compare VLS measurements with a validated quantitative microvascular oxygen tension (μPO) measurement technique.

Methods: Simultaneous VLS measurements and μPO measurements were performed on the small intestine of five pigs.

Characterization of the flavin monooxygenase involved in biosynthesis of the antimalarial FR-900098.

The latter steps in this biosynthetic pathway for the antimalarial phosphonic acid FR-900098 include the installation of a hydroxamate onto 3-aminopropylphosphonate, which is catalyzed by the consecutive actions of an acetyltransferase and an amine hydroxylase. Here, we present the 1.6 Å resolution co-crystal structure and accompanying biochemical characterization of FrbG, which catalyzes the hydroxylation of aminopropylphosphonate.

A monitor for Cellular Oxygen METabolism (COMET): monitoring tissue oxygenation at the mitochondrial level.

After introduction of the protoporphyrin IX-triplet state lifetime technique as a new method to measure mitochondrial oxygen tension in vivo, the development of a clinical monitor was started. This monitor is the "COMET", an acronym for Cellular Oxygen METabolism. The COMET is a non-invasive electrically powered optical device that allows measurements on the skin.

Cutaneous Mitochondrial PO2, but Not Tissue Oxygen Saturation, Is an Early Indicator of the Physiologic Limit of Hemodilution in the Pig.

Background: Hemodilution is a consequence of fluid replacement during blood loss and is limited by the individual ability to compensate for decreasing hemoglobin level. We tested the ability of a novel noninvasive method for measuring cutaneous mitochondrial PO2 (mitoPO2) to detect this threshold early.

Methods: Anesthetized and ventilated pigs were hemodynamically monitored and randomized into a hemodilution (n = 12) or a time control (TC) group (n = 14).

Activated protein C ameliorates impaired renal microvascular oxygenation and sodium reabsorption in endotoxemic rats.

Introduction: We aimed to test whether continuous recombinant human activated protein C (APC) administration would be able to protect renal oxygenation and function during endotoxemia in order to provide more insight into the role of coagulation and inflammation in the development of septic acute kidney injury.

Methods: In anesthetized, mechanically ventilated Wistar rats, endotoxemia was induced by lipopolysaccharide administration (10 mg/kg i.v.

Method for assembling and expressing multiple genes in the nucleus of microalgae.

The green alga, Chlamydomonas reinhardtii, is a model organism used in the study of photosynthesis and biotechnological research. Despite its importance, a complete set of genetic tools has yet to be developed. Here, we report the development of a new method for constructing a multi-gene pathway in Saccharomyces cerevisiae and integrating the assembled pathway into the nuclear genome of C.

Acute normovolemic hemodilution in the pig is associated with renal tissue edema, impaired renal microvascular oxygenation, and functional loss.

Background: The authors investigated the impact of acute normovolemic hemodilution (ANH) on intrarenal oxygenation and its functional short-term consequences in pigs.

Methods: Renal microvascular oxygenation (µPO2) was measured in cortex, outer and inner medulla via three implanted optical fibers by oxygen-dependent quenching of phosphorescence. Besides systemic hemodynamics, renal function, histopathology, and hypoxia-inducible factor-1α expression were determined.

Production of xylitol by recombinant microalgae.

Microalgae have received significant attention recently as a potential low-cost host for the production of next-generation biofuels and natural products. Here we show that the chloroplast genome of the eukaryotic green microalga Chlamydomonas reinhardtii can be genetically engineered to produce xylitol through the introduction of a gene encoding a xylose reductase (XR) from the fungi Neurospora crassa. Increased levels of heterologous protein accumulation and xylitol production were achieved by synthesizing the XR gene in the chloroplast codon bias and by driving expression of the codon-optimized XR gene using a 16S/atpA promoter/5'-UTR fusion.

Method to assemble and integrate biochemical pathways into the chloroplast genome of Chlamydomonas reinhardtii.

Recombinant protein expression in the chloroplasts of green algae has recently become more routine; however, the heterologous expression of multiple proteins or complete biosynthetic pathways remains a significant challenge. Here, we show that a modified DNA Assembler approach can be used to rapidly assemble multiple-gene biosynthetic pathways in yeast and then integrate these assembled pathways at a site-specific location in the chloroplast genome of the microalgal species Chlamydomonas reinhardtii. As a proof of concept, this method was used to successfully integrate and functionally express up to three reporter proteins (AphA6, AadA, and GFP) in the chloroplast of C.

Crystal structures of phosphite dehydrogenase provide insights into nicotinamide cofactor regeneration.

The enzyme phosphite dehydrogenase (PTDH) catalyzes the NAD(+)-dependent conversion of phosphite to phosphate and represents the first biological catalyst that has been shown to conduct the enzymatic oxidation of phosphorus. Despite investigation for more than a decade into both the mechanism of its unusual reaction and its utility in cofactor regeneration, there has been a lack of any structural data for PTDH. Here we present the cocrystal structure of an engineered thermostable variant of PTDH bound to NAD(+) (1.

Investigation of the role of Arg301 identified in the X-ray structure of phosphite dehydrogenase.

Phosphite dehydrogenase (PTDH) from Pseudomonas stutzeri catalyzes the nicotinamide adenine dinucleotide-dependent oxidation of phosphite to phosphate. The enzyme belongs to the family of D-hydroxy acid dehydrogenases (DHDHs). A search of the protein databases uncovered many additional putative phosphite dehydrogenases.

Effects of different leukocyte subpopulations and flow conditions on leukocyte accumulation during reperfusion.

Background/aims: The study examined the interdependent effects of shear stress and different leukocyte subpopulations on endothelial cell activation and cell interactions during low flow and reperfusion.

Methods: Human umbilical venous endothelial cells were perfused with either neutrophils or monocytes at different shear stress (2-0.25 dyn/cm(2)) and adhesion was quantified by microscopy.

Directed evolution: selection of the host organism.

Directed evolution has become a well-established tool for improving proteins and biological systems. A critical aspect of directed evolution is the selection of a suitable host organism for achieving functional expression of the target gene. To date, most directed evolution studies have used either Escherichia coli or Saccharomyces cerevisiae as a host; however, other bacterial and yeast species, as well as mammalian and insect cell lines, have also been successfully used.

Microvascular and mitochondrial PO(2) simultaneously measured by oxygen-dependent delayed luminescence.

Measurement of tissue oxygenation is a complex task and various techniques have led to a wide range of tissue PO(2) values and contradictory results. Tissue is compartmentalized in microcirculation, interstitium and intracellular space and current techniques are biased towards a certain compartment. Simultaneous oxygen measurements in various compartments might be of great benefit for our understanding of determinants of tissue oxygenation.

Oxygen-dependent delayed fluorescence measured in skin after topical application of 5-aminolevulinic acid.

Mitochondrial oxygen tension can be measured in vivo by means of oxygen-dependent quenching of delayed fluorescence of protoporphyrin IX (PpIX). Here we demonstrate that delayed fluorescence is readily observed from skin in rat and man after topical application of the PpIX precursor 5-aminolevulinic acid (ALA). Delayed fluorescence lifetimes respond to changes in inspired oxygen fraction and blood supply.

Deciphering the late biosynthetic steps of antimalarial compound FR-900098.

FR-900098 is a potent chemotherapeutic agent for the treatment of malaria. Here we report the heterologous production of this compound in Escherichia coli by reconstructing the entire biosynthetic pathway using a three-plasmid system. Based on this system, whole-cell feeding assays in combination with in vitro enzymatic activity assays reveal an unusual functional role of nucleotide conjugation and lead to the complete elucidation of the previously unassigned late biosynthetic steps.

Cloning and heterologous expression of the spectinabilin biosynthetic gene cluster from Streptomyces spectabilis.

Spectinabilin is a rare nitrophenyl-substituted polyketide metabolite. Here we report the cloning and heterologous expression of the spectinabilin gene cluster from Streptomyces spectabilis. Unexpectedly, this gene cluster is evolutionarily closer to the aureothin gene cluster than to the spectinabilin gene cluster from Streptomyces orinoci.