An Image Segmentation by BFV and TLBO

This paper presents the establishing of a biconvex fuzzy variational (BFV) method with teaching learning based optimization (TLBO) for geometric image segmentation (GIS). Firstly, a biconvex object function is adopted to process GIS. Then, TLBO is introduced to maximally optimize the length penalty item (LPI), which will be changed under teaching and learner phase of TLBO, making the LPI closer to the target boundary. Afterward, the LPI can be adjusted based on fitness function, namely, the evaluation standards of image quality. Finally, the LP is combined item with the numerical order to get better results. Different GIS strategies are compared with various fitness functions in terms of accuracy. Simulations show that the presented method is more effective in this area

Current Frontiers and Perspectives in Cell Biology

A numerous internationally renowned authors in the pages of this book present the views of the fields of cell biology and their own research results or review of current knowledge. Chapters are divided into five sections that are dedicated to cell structures and functions, genetic material, regulatory mechanisms, cellular biomedicine and new methods in cell biology. Multidisciplinary and often quite versatile approach by many authors have imposed restrictions of this classification, so it is certain that many chapters could belong to the other sections of this book. The current frontiers, on the manner in which they described in the book, can be a good inspiration to many readers for further improving, and perspectives which are highlighted can be seen in many areas of fundamental biology, biomedicine, biotechnology and other applications of knowledge of cell biology. The book will be very useful for beginners to gain insight into new area, as well as experts to find new facts and expanding horizons

27th Fungal Genetics Conference

Program and abstracts from the 27th Fungal Genetics Conference Asilomar, March 12-17, 2013

27th Fungal Genetics Conference

Program and abstracts from the 27th Fungal Genetics Conference Asilomar, March 12-17, 2013

[¹⁸F]fluorination of biorelevant arylboronic acid pinacol ester scaffolds synthesized by convergence techniques

Aim: The development of small molecules through convergent multicomponent reactions (MCR) has been boosted during the last decade due to the ability to synthesize, virtually without any side-products, numerous small drug-like molecules with several degrees of structural diversity.(1) The association of positron emission tomography (PET) labeling techniques in line with the “one-pot” development of biologically active compounds has the potential to become relevant not only for the evaluation and characterization of those MCR products through molecular imaging, but also to increase the library of radiotracers available. Therefore, since the [18F]fluorination of arylboronic acid pinacol ester derivatives tolerates electron-poor and electro-rich arenes and various functional groups,(2) the main goal of this research work was to achieve the 18F-radiolabeling of several different molecules synthesized through MCR. Materials and Methods: [18F]Fluorination of boronic acid pinacol esters was first extensively optimized using a benzaldehyde derivative in relation to the ideal amount of Cu(II) catalyst and precursor to be used, as well as the reaction solvent. Radiochemical conversion (RCC) yields were assessed by TLC-SG. The optimized radiolabeling conditions were subsequently applied to several structurally different MCR scaffolds comprising biologically relevant pharmacophores (e.g. β-lactam, morpholine, tetrazole, oxazole) that were synthesized to specifically contain a boronic acid pinacol ester group. Results: Radiolabeling with fluorine-18 was achieved with volumes (800 μl) and activities (≤ 2 GBq) compatible with most radiochemistry techniques and modules. In summary, an increase in the quantities of precursor or Cu(II) catalyst lead to higher conversion yields. An optimal amount of precursor (0.06 mmol) and Cu(OTf)2(py)4 (0.04 mmol) was defined for further reactions, with DMA being a preferential solvent over DMF. RCC yields from 15% to 76%, depending on the scaffold, were reproducibly achieved. Interestingly, it was noticed that the structure of the scaffolds, beyond the arylboronic acid, exerts some influence in the final RCC, with electron-withdrawing groups in the para position apparently enhancing the radiolabeling yield. Conclusion: The developed method with high RCC and reproducibility has the potential to be applied in line with MCR and also has a possibility to be incorporated in a later stage of this convergent “one-pot” synthesis strategy. Further studies are currently ongoing to apply this radiolabeling concept to fluorine-containing approved drugs whose boronic acid pinacol ester precursors can be synthesized through MCR (e.g. atorvastatin)

Cellular and molecular mechanisms of arrhythmias in cardiac fibrosis and beyond : from symptoms to substrates towards solutions

Currently, treatment of arrhythmias is largely symptomatic and pro-arrhythmic mechanisms are incompletely understood. The research described in this thesis therefore investigates cellular and molecular pro-arrhythmic mechanisms in in vitro models of pro-arrhythmic substrates such as fibrosis and hypertrophy, to provide and expand upon a mechanistic basis for future, substrate-oriented anti-arrhythmic strategies. In the model of cardiac fibrosis, the prominent pro-arrhythmic role of myofibroblasts was apparent. In a quantity-dependent manner, these cells have a detrimental depolarizing and pro-arhythmic influence on cardiac tissue. A mechanism that appears to be responsible for this depolarization of cardiomyocytes is heterocellular coupling, a pro-arrhythmic mechanism that was also observed between mesenchymal stem cells and cardiomyocytes and may therefore also have cautionary implications for the future of cardiac stem cell therapy. Through the use of genetic modification, such mechanisms can be selectively targeted. In this thesis, control of selective transgene expression in myofibroblasts or cardiomyocytes was further refined by investigating the cellular tropism of several adeno-associated viral vectors and the use of celltype-specific promotors. By employing such genetic tools together with expanded knowledge of pro-arrhythmic mechanisms of cardiac fibrosis and other pro-arrhythmic substrates, future treatment modalities for arrhythmias may improve by becoming more mechanism- and substrate-oriented.Dutch Heart Foundation (2008/B119)for conducted studies. Scimedia and Greiner Bio-One for the publication of the thesisUBL - phd migration 201

Editing the genome of chicken primordial germ cells to introduce alleles and study gene function

With continuing advances in genome sequencing technology, the chicken genome assembly is now better annotated with improved accuracy to the level of single nucleotide polymorphisms. Additionally, the genomes of other birds such as the duck, turkey and zebra finch have now been sequenced. A great opportunity exists in avian biology to use genome editing technology to introduce small and defined sequence changes to create specific haplotypes in chicken to investigate gene regulatory function, and also perform rapid and seamless transfer of specific alleles between chicken breeds. The methods for performing such precise genome editing are well established for mammalian species but are not readily applicable in birds due to evolutionary differences in reproductive biology. A significant leap forward to address this challenge in avian biology was the development of long-term culture methods for chicken primordial germ cells (PGCs). PGCs present a cell line in which to perform targeted genetic manipulations that will be heritable. Chicken PGCs have been successfully targeted to generate genetically modified chickens. However, genome editing to introduce small and defined sequence changes has not been demonstrated in any avian species. To address this deficit, the application of CRISPR/Cas9 and short oligonucleotide donors in chicken PGCs for performing small and defined sequence changes was investigated in this thesis. Specifically, homology-directed DNA repair (HDR) using oligonucleotide donors along with wild-type CRISPR/Cas9 (SpCas9-WT) or high fidelity CRISPR/Cas9 (SpCas9-HF1) was investigated in cultured chicken PGCs. The results obtained showed that small sequences changes ranging from a single to a few nucleotides could be precisely edited in many loci in chicken PGCs. In comparison to SpCas9-WT, SpCas9-HF1 increased the frequency of biallelic and single allele editing to generate specific homozygous and heterozygous genotypes. This finding demonstrates the utility of high fidelity CRISPR/Cas9 variants for performing sequence editing with high efficiency in PGCs. Since PGCs can be converted into pluripotent stem cells that can potentially differentiate into many cell types from the three germ layers, genome editing of PGCs can, therefore, be used to generate PGC-derived avian cell types with defined genetic alterations to investigate the host-pathogen interactions of infectious avian diseases. To investigate this possibility, the chicken ANP32A gene was investigated as a target for genetic resistance to avian influenza virus in PGC-derived chicken cell lines. Targeted modification of ANP32A was performed to generate clonal lines of genome-edited PGCs. Avian influenza minigenome replication assays were subsequently performed in the ANP32A-mutant PGC-derived cell lines. The results verified that ANP32A function is crucial for the function of both avian virus polymerase and human-adapted virus polymerase in chicken cells. Importantly, an asparagine to isoleucine mutation at position 129 (N129I) in chicken ANP32A failed to support avian influenza polymerase function. This genetic change can be introduced into chickens and validated in virological studies. Importantly, the results of my investigation demonstrate the potential to use genome editing of PGCs as an approach to generate many types of unique cell models for the study of avian biology. Genome editing of PGCs may also be applied to unravel the genes that control the development of the avian germ cell lineage. In the mouse, gene targeting has been extensively applied to generate loss-of-function mouse models to use the reverse genetics approach to identify key genes that regulate the migration of specified PGCs to the genital ridges. Avian PGCs express similar cytokine receptors as their mammalian counterparts. However, the factors guiding the migration of avian PGCs are largely unknown. To address this, CRISPR/Cas9 was used in this thesis to generate clonal lines of chicken PGCs with loss-of-function deletions in the CXCR4 and c-Kit genes which have been implicated in controlling mouse PGC migration. The results showed that CXCR4-deficient PGCs are absent from the gonads whereas c-Kit-deficient PGCs colonise the developing gonads in reduced numbers and are significantly reduced or absent from older stages. This finding shows a conserved role for CXCR4 and c-Kit signalling in chicken PGC development. Importantly, other genes suspected to be involved in controlling the development of avian germ cells can be investigated using this approach to increase our understanding of avian reproductive biology. Finally, the methods developed in this thesis for editing of the chicken genome may be applied in other avian species once culture methods for the PGCs from these species are develope

Biotechnology and Crop Improvement in Asia

This book results from a workshop held at ICRISAT 3-7 Dec 1990 by the Asian Development Bank (ADB). The participants were representatives of Asian countries, and scientific leaders in [he various fields of biotechnology The introductory chapter outlines the role of the ADS in strengthening biotechnology research in Asia; this is followed by a presentation of the Indian biotechnology program and the problems thaI are being addressed. A chapter on Industry and Public Sector the and of private companics becoming involved in biotechnology research. The country representatives presented papers on the slatus of biotechnology in ,heir country, and these chaplers give an indication of the wide range of achievement in the area. Participants also outlined those crops and areas of crop improvement to which biotechnology could he applied. The remaining are reviews of the major disciplines in biotechnology and reports by Asian scientists and others on research relevant to Asian agriculture. These reviews cover cell lind tissue culture. with emphasis on cereals. and haploids. There are four reviews on transformation, covering Agrobacterium-medialed transformation, physical methods gene transfer the use of viruses as vectors and gene action in transformed plants. The section on genome characterization and diagnostics covers the genomes of plants-nuclear mitochondrial, and plastid and of viruses, and the relevance of genome research to plant improvement, through the use of markers-restriction fragment length polymorph isms (RFLP), polymerase chain reaclion (PCR), and random amplified polymorphic DNA (RAPD). The recommendations of the workshop give a useful insight into how a representative cross section view the future of biotechnology. Title book contains a glossary and index