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in vitro evolution

Kento Tokuyama, Yoshihiro Toya, Takaaki Horinouchi, Chikara Furusawa, Fumio Matsuda, Hiroshi Shimizu
Gene deletion strategies using flux balance analysis (FBA) have improved the growth-coupled production of various compounds. However, the productivities were often below the expectation because the cells failed to adapt to these genetic perturbations. Here, we demonstrate the productivity of the succinate of the designed gene deletion strain was improved by adaptive laboratory evolution (ALE). Although FBA predicted deletions of adhE-pykAF-gldA-pflB lead to produce succinate from glycerol with a yield of 0...
February 19, 2018: Biotechnology and Bioengineering
Madeline R Luth, Purva Gupta, Sabine Ottilie, Elizabeth A Winzeler
Although many new anti-infectives have been discovered and developed solely using phenotypic cellular screening and assay optimization, most researchers recognize that structure-guided drug design is more practical and less costly. In addition, a greater chemical space can be interrogated with structure-guided drug design. The practicality of structure-guided drug design has launched a search for the targets of compounds discovered in phenotypic screens. One method that has been used extensively in malaria parasites for target discovery and chemical validation is in vitro evolution and whole genome analysis (IVIEWGA)...
February 16, 2018: ACS Infectious Diseases
Sylvin Benjamin Ateba, Sadrine Tchoukouegno Ngeu, Marie Alfrede Mvondo, Job Tchoumtchoua, Charline Florence Awounfack, Liselotte Krenn, Dieudonne Njamen
BACKGROUND: The approval of Taxol® in 1993 marked the great entrance of terpenoids in the anti-cancer area and this drug is still highly important in the treatment of refractory ovarian, breast and other cancers. Over decades, other prominent natural terpenoids have become indispensable for the modern pharmacotherapy of breast cancer. However, given the rapid evolution of drug resistance, effective treatments for advanced breast cancers requiring cytotoxic chemotherapy represent a major unmet clinical need...
February 13, 2018: Current Medicinal Chemistry
Zhihao Tan, De Zhi Valerie Chu, Yong Jie Andrew Chan, Yi Ena Lu, Giulia Rancati
Polyploidization, a common event during the evolution of different tumours, has been proposed to confer selective advantages to tumour cells by increasing the occurrence of mutations promoting cancer progression and by conferring chemotherapy resistance. While conditions leading to polyploidy in cancer cells have been described, a general mechanism explaining the incidence of this karyotypic change in tumours is still missing. In this study, we tested whether a widespread tumour microenvironmental condition, low pH, could induce polyploidization in mammalian cells...
February 13, 2018: Scientific Reports
Dominik Schwarz, Orit Adato, Amnon Horovitz, Ron Unger
The GroE chaperonin system, which comprises GroEL and GroES, assists protein folding in vivo and in vitro. It is conserved in all prokaryotes except in most, but not all, members of the class of mollicutes. In Escherichia coli, about 60 proteins were found to be obligatory clients of the GroE system. Here, we describe the properties of the homologs of these GroE clients in mollicutes and the evolution of chaperonins in this class of bacteria. Comparing the properties of these homologs in mollicutes with and without chaperonins enabled us to search for features correlated with the presence of GroE...
2018: PloS One
Anthony R Scialli, George Daston, Connie Chen, Prägati S Coder, Susan Y Euling, Jennifer Foreman, Alan M Hoberman, Julia Hui, Thomas Knudsen, Susan L Makris, LaRonda Morford, Aldert H Piersma, Dinesh Stanislaus, Kary E Thompson
BACKGROUND: Current developmental toxicity testing adheres largely to protocols suggested in 1966 involving the administration of test compound to pregnant laboratory animals. After more than 50 years of embryo-fetal development testing, are we ready to consider a different approach to human developmental toxicity testing? METHODS: A workshop was held under the auspices of the Developmental and Reproductive Toxicology Technical Committee of the ILSI Health and Environmental Sciences Institute to consider how we might design developmental toxicity testing if we started over with 21st century knowledge and techniques (revolution)...
February 12, 2018: Birth Defects Research
Shao-Yao Ying, Donald C Chang, Shi-Lung Lin
MicroRNAs (miRNAs), widely distributed, small regulatory RNA genes, target both messenger RNA (mRNA) degradation and suppression of protein translation based on sequence complementarity between the miRNA and its targeted mRNA. Different names have been used to describe various types of miRNA. During evolution, RNA retroviruses or transgenes invaded the eukaryotic genome and were inserted itself in the noncoding regions of DNA, conceivably acting as transposon-like jumping genes, providing defense from viral invasion and fine-tuning of gene expression as a secondary level of gene modulation in eukaryotes...
2018: Methods in Molecular Biology
Yingchi Zhang, Yufeng Gao, Hui Zhang, Jingliao Zhang, Fuhong He, Aleš Hnízda, Maoxiang Qian, Xiaoming Liu, Yoshihiro Gocho, Ching-Hon Pui, Tao Cheng, Qianfei Wang, Jun J Yang, Xiaofan Zhu, Xin Liu
Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) comprises of approximately 10-15% of childhood ALL cases, many of whom respond exquisitely to tyrosine kinase inhibitors (TKIs), e.g., imatinib in PDGFRB -rearranged ALL. However, some cases developed drug resistance to TKIs with mechanisms poorly understood. In this study, we identified a novel PDGFRB fusion gene, namely AGGF1-PDGFRB , and functionally characterized its oncogenic potential in vitro. Further genomic profiling of longitudinally collected samples during treatment revealed the emergence of a mutation PDGFRBC843G , which directly conferred resistance to all generations of ABL TKIs, including imatinib, dasatinib, nilotinib, and ponatinib...
February 6, 2018: Blood
Prabh Basra, Ahlam Alsaadi, Gabriel Bernal-Astrain, Michael Liam O'Sullivan, Bryn Hazlett, Leah Marie Clarke, Andrew Schoenrock, Sylvain Pitre, Alex Wong
Evolutionary trade-offs occur when selection on one trait has detrimental effects on other traits. In pathogenic microbes, it has been hypothesized that antibiotic resistance trades off with fitness in the absence of antibiotic. While studies of single resistance mutations support this hypothesis, it is unclear whether trade-offs are maintained over time, due to compensatory evolution and broader effects of genetic background. Here, we leverage natural variation in 39 extra-intestinal clinical isolates of Escherichia coli to assess trade-offs between growth rates and resistance to fluoroquinolone and cephalosporin antibiotics...
February 7, 2018: Genome Biology and Evolution
Margriet M Palm, Marjet Elemans, Joost B Beltman
Tumors consist of a hierarchical population of cells that differ in their phenotype and genotype. This hierarchical organization of cells means that a few clones (i.e., cells and several generations of offspring) are abundant while most are rare, which is called clonal dominance. Such dominance also occurred in published in vitro iterated growth and passage experiments with tumor cells in which genetic barcodes were used for lineage tracing. A potential source for such heterogeneity is that dominant clones derive from cancer stem cells with an unlimited self-renewal capacity...
February 12, 2018: PLoS Computational Biology
Jyoti Bala, Srinivasan Chinnapaiyan, Rajib Kumar Dutta, Hoshang Unwalla
Aptamers are high affinity single-stranded nucleic acid or protein ligands which exhibit specificity and avidity comparable to, or exceeding that of antibodies and can be generated against most targets. The functionality of aptamers is based on their unique tertiary structure, complexity and their ability to attain unique binding pockets by folding. Aptamers are selected in vitro by a process called Systematic Evolution of Ligands by Exponential enrichment (SELEX). The Kd values for the selected aptamer are often in the picomolar to low nanomolar range...
February 12, 2018: RNA Biology
Céline Engrand, Didier Laux, Jean-Yves Ferrandis, Roland Demaria, Emmanuel Le Clézio
Tissue temperature control during cardiac surgery is crucial for myocardial protection. To preserve the tissue, a hypothermic cardioplegia is applied in order to decrease the heart temperature down to around 10°C. The monitoring of the thermal evolution of the myocardium is then of importance to minimize deleterious effects on the heart. The present work aims at evaluating the potential of an ultrasonic velocimetric thermometry on the monitoring of in vitro tissues heating. An indentation process is first proposed to identify the experimental linear relationship linking, in myocardia, the speed of the ultrasonic longitudinal wave to the tissue temperature...
February 1, 2018: Ultrasonics
Shin-Hye Park, Takanori Uzawa, Fumiyuki Hattori, Shuichi Ogino, Naoki Morimoto, Satoshi Tsuneda, Yoshihiro Ito
To enhance the therapeutic effect of growth factors, a powerful strategy is to direct their localization to damaged sites. To treat skin wounds and myocardial infarction, we selected vascular endothelial growth factor (VEGF) carrying binding affinity to collagen. A simple conjugation of a reported collagen-binding sequence and VEGF did not increase the collagen-binding affinity, indicating that the molecular interaction between the two proteins abolished collagen binding activity. Here, we present a new molecular evolution strategy, "all-in-one" in vitro selection, in which a collagen-binding VEGF (CB-VEGF) was directly identified from a random library consisting of random and VEGF sequences...
February 3, 2018: Biomaterials
Lei Chen, Sheng Chen, Jing Wu, Dan Wu
We used in vitro molecular evolution technology by error-prone PCR and high-throughput screening to improve thermostability of Bacillus flexus CCTCC 2015368 β-amylase. Mutant D476N with significant thermostability increase was selected by LB agar starch plate colorimetric assay and 96-well plate enzyme activity assay. The optimum pH was 6.5 for the mutant D476N, compared to 7.0 of the wild type. The optimal temperature was 55 ℃ for both mutant D476N and the wild type. The T₅₀ value of the mutant D476N was 4 ℃ higher than that of the wild type...
February 25, 2018: Sheng Wu Gong Cheng Xue Bao, Chinese Journal of Biotechnology
Johanna Zemva, Daniel Pfaff, Jan B Groener, Thomas Fleming, Stephan Herzig, Aurelio Teleman, Peter P Nawroth, Jens Tyedmers
Levels of reactive metabolites such as reactive carbonyl and oxygen species are increased in patients with diabetes mellitus. The most important reactive dicarbonyl species, methylglyoxal (MG), formed as by-product during glucose metabolism, is more and more recognized as a trigger for the development and progression of diabetic complications. Although it is clear that MG provokes toxic effects, it is currently not well understood what cellular changes MG induces on a molecular level that may lead to pathophysiological conditions found in long-term diabetic complications...
February 8, 2018: Experimental and Clinical Endocrinology & Diabetes
Yuki Ogura-Tsujita, Gerhard Gebauer, Hui Xu, Yu Fukasawa, Hidetaka Umata, Kenshi Tetsuka, Miho Kubota, Julienne M-I Schweiger, Satoshi Yamashita, Nitaro Maekawa, Masayuki Maki, Shiro Isshiki, Tomohisa Yukawa
The climbing orchid Erythrorchis altissima is the largest mycoheterotroph in the world. Although previous in vitro work suggests that E. altissima has a unique symbiosis with wood-decaying fungi, little is known about how this giant orchid meets its carbon and nutrient demands exclusively via mycorrhizal fungi. In this study, the mycorrhizal fungi of E. altissima were molecularly identified using root samples from 26 individuals. Furthermore, in vitro symbiotic germination with five fungi and stable isotope compositions in five E...
February 8, 2018: Molecular Ecology
Adam Barlev, Dipankar Sen
Photochemical modification is the major class of environmental damage suffered by DNA, the genetic material of all free-living organisms. Photolyases are enzymes that carry out direct photochemical repair (photoreactivation) of covalent pyrimidine dimers formed in DNA from exposure to ultraviolet light. The discovery of catalytic RNAs in the 1980s led to the "RNA world hypothesis", which posits that early in evolution RNA or a similar polymer served both genetic and catalytic functions. Intrigued by the RNA world hypothesis, we set out to test whether a catalytic RNA (or a surrogate, a catalytic DNA) with photolyase activity could be contemplated...
February 8, 2018: Accounts of Chemical Research
Damarys Relova, Liliam Rios, Ana M Acevedo, Liani Coronado, Carmen L Perera, Lester J Pérez
The current global conditions, which include intensive globalization, climate changes, and viral evolution among other factors, have led to an increased emergence of viruses and new viral diseases; RNA viruses are key drivers of this evolution. Laboratory networks that are linked to central reference laboratories are required to conduct both active and passive environmental surveillance of this complicated global viral environment. These tasks require a continuous exchange of strains or field samples between different diagnostic laboratories...
February 6, 2018: Veterinary Sciences
Sebastian Schneider, Michael Rudolph, Vanessa Bause, Andreas Terfort
The infection of dental implants may cause severe inflammation of tissue and even bone degradation if not treated. For titanium implants, a new, minimally invasive approach is the electrochemical removal of the biofilms including the disinfection of the metal surface. In this project, several parameters, such as electrode potentials and electrolyte compositions, were varied to understand the underlying mechanisms. Optimal electrolytes contained iodide as well as lactic acid. Electrochemical experiments, such as cyclic voltammetry or measurements of open circuit potentials, were performed in different cell set-ups to distinguish between different possible reactions...
January 29, 2018: Bioelectrochemistry
Hai-Ming Wang, Tian-Xin Liu, Tong-Yun Wang, Gang Wang, Yong-Gang Liu, Si-Guo Liu, Yan-Dong Tang, Xue-Hui Cai
Porcine reproductive and respiratory syndrome virus (PRRSV) is a pathogen of great economic significance that impacts the swine industry globally. Since the first report of a porcine reproductive and respiratory syndrome (PRRS) outbreak, tremendous efforts to control this disease, including various national policies and plans incorporating the use of multiple modified live-virus vaccines, have been made. However, PRRSV is still a significant threat to the swine industry, and new variants continually emerge as a result of PRRSV evolution...
February 6, 2018: Archives of Virology
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