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whole mount in situ hybridization

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https://www.readbyqxmd.com/read/29203198/phylogeny-and-expression-patterns-of-two-apolipoprotein-e-genes-in-the-flatfish-senegalese-sole
#1
Javier Roman-Padilla, Ana Rodríguez-Rúa, Carlos Carballo, Manuel Manchado, Ismael Hachero-Cruzado
The apolipoprotein E (ApoE) is a key component of several lipoproteins involved in lipid homeostasis. In this study, two cDNA sequences encoding ApoE (referred to as apoEa and apoEb) were characterized in the flatfish Solea senegalensis. The predicted peptides contained conserved structural blocks related with their capacity for lipid binding and lipoprotein receptor interaction. At genomic level, both genes contained five exons and four introns and they were organized into two tandem arrays with apoA-IV gene copies...
December 1, 2017: Gene
https://www.readbyqxmd.com/read/29196253/involvement-of-lypge-in-the-formation-of-eye-and-pineal-gland-in-zebrafish
#2
Dongrui Ji, Su Wang, Mingyue Li, Shicui Zhang, Hongyan Li
The proteins of Ly-6 (lymphocyte antigen-6) family are involved in the regulation of immunoreaction, cell migration and adhesion, and neuronal excitability. However, little is known about the function of Ly-6 proteins in embryogenesis. Herein, we identified a GPI anchored Ly-6 member named ly6 expressed in pineal gland and eye (lypge). Dynamic expression pattern of lypge was revealed by whole mount in situ hybridization. It was strikingly expressed in the pineal gland and cone photoreceptor, and its expression was regulated by orthodenticle homolog 5 (otx5) which has been shown to control the expression of many pineal genes...
November 28, 2017: Gene
https://www.readbyqxmd.com/read/29130196/super-resolution-single-molecule-fish-at-the-drosophila-neuromuscular-junction
#3
Joshua S Titlow, Lu Yang, Richard M Parton, Ana Palanca, Ilan Davis
The lack of an effective, simple, and highly sensitive protocol for fluorescent in situ hybridization (FISH) at the Drosophila larval neuromuscular junction (NMJ) has hampered the study of mRNA biology. Here, we describe our modified single molecule FISH (smFISH) methods that work well in whole mount Drosophila NMJ preparations to quantify primary transcription and count individual cytoplasmic mRNA molecules in specimens while maintaining ultrastructural preservation. The smFISH method is suitable for high-throughput sample processing and 3D image acquisition using any conventional microscopy imaging modality and is compatible with the use of antibody colabeling and transgenic fluorescent protein tags in axons, glia, synapses, and muscle cells...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29075432/establishment-of-molecular-genetic-approaches-to-study-gene-expression-and-function-in-an-invasive-hemipteran-halyomorpha-halys
#4
Yong Lu, Mengyao Chen, Katie Reding, Leslie Pick
Hemiptera is a large clade of insects understudied in terms of developmental biology. Halyomorpha halys, the Brown Marmorated Stink Bug (BMSB, referred to throughout as H. halys), is an invasive hemipteran pest of the mid-Atlantic region of the USA that has rapidly spread to other regions in recent years, devastating a wide range of crops using a piercing and sucking mechanism. Its phylogenetic position, polyphagous habits, and rapid spread in the USA suggested that H. halys would be an ideal system to broaden our knowledge of developmental mechanisms in insects...
2017: EvoDevo
https://www.readbyqxmd.com/read/29052208/fluorescence-in-situ-hybridization-fish-and-immunolabeling-on-3d-preserved-nuclei
#5
Till David Bey, Maria Koini, Paul Fransz
The spatial distribution of genes in the nucleus emerges as an important factor in gene regulation and epigenetics. The position of loci relative to each other, to nuclear landmarks such as the nucleolus and chromocenters, as well as to chromatin proteins is therefore highly interesting. With fluorescent in situ hybridization (FISH) specific DNA sequences can be stained and antibodies allow the detection of specific proteins. Here, we present two protocols that preserve the 3D structure of nuclei. With whole-mount FISH, specific sequences can be stained in intact tissues and, secondly, a combined immunolabeling and FISH protocol on acrylamide-embedded nuclei makes it possible to stain DNA sequences and proteins simultaneously...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29030842/microrna-detection-by-whole-mount-in-situ-hybridization-in-c-elegans
#6
Yoshiki Andachi, Yuji Kohara
MicroRNAs (miRNAs) loaded on argonaute proteins guide RNA-induced silencing complexes to target mRNAs. An excellent method to decipher the spatiotemporal expression patterns of miRNAs is whole-mount in situ hybridization (WISH), which has been successfully used in vertebrate embryos but still remains unavailable for many animal species. Here, we describe a WISH method for miRNA detection in Caenorhabditis elegans at both embryonic and post-embryonic stages. Strategies devised for detection include fixation of animals with carbodiimide at a high temperature and subsequent partial digestion of the fixed animals with an extremely high concentration of proteinase...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29024245/hace1-influences-zebrafish-cardiac-development-via-ros-dependent-mechanisms
#7
Babak Razaghi, Shelby L Steele, Sergey V Prykhozhij, Matthew R Stoyek, Jessica A Hill, Matthew D Cooper, Lindsay Mcdonald, William Lin, Mads Daugaard, Nicolas Crapoulet, Simi Chacko, Stephen M Lewis, Ian C Scott, Poul H B Sorensen, Jason N Berman
BACKGROUND: In this study, we reveal a previously undescribed role of the HACE1 (HECT domain and Ankyrin repeat Containing E3 ubiquitin-protein ligase 1) tumor suppressor protein in normal vertebrate heart development using the zebrafish (Danio rerio) model. We examined the link between the cardiac phenotypes associated with hace1 loss of function to the expression of the Rho small family GTPase, rac1, which is a known target of HACE1 and promotes ROS production via its interaction with NADPH oxidase holoenzymes...
October 11, 2017: Developmental Dynamics: An Official Publication of the American Association of Anatomists
https://www.readbyqxmd.com/read/28987646/molecular-cloning-and-expression-of-octamer-binding-transcription-factor-oct4-in-the-large-yellow-croaker-larimichthys-crocea
#8
Yonghua Jiang, Kunhuang Han, Shihai Chen, Mingyi Cai, Yilei Wang, Ziping Zhang
Octamer-binding transcription factor 4 (Oct4) is a crucial pluripotent transcription factor in controlling pluripotency of embryonic stem cells (ESCs), formation of primordial germ cells (PGCs), early embryonic development, and gonadal germ cells. To understand the roles of Oct4 in the large yellow croaker (Larimichthys crocea) (Lc-Oct4), the full-length cDNA of Oct4 was cloned and analyzed. Lc-Oct4 includes a 104-bp 5' untranslated region (UTR), a 567-bp 3' UTR and a 1431-bp coding region encoding a protein of 476 amino acids (aa) with a predicted molecular mass (Mm) of 52...
October 4, 2017: Gene Expression Patterns: GEP
https://www.readbyqxmd.com/read/28964895/functions-of-the-vasa-gene-in-schistosoma-japonicum-as-assessed-by-rna-interference
#9
Siyu He, Lulu Zhu, Fengchun Liu, Quan Liu, Yanjing Shao, Mengqing Hua, Han Ding, Wei Shao, Yinan Du, Xin Hou, Cuiping Ren, Miao Liu, Jijia Shen
Vasa, an enzyme belonging to the helicase family, contributes to the regulation of reproductive system development in many species. Thus, we hypothesized that the Vasa3 gene may function in the reproductive system of the parasite Schistosoma japonicum (S. japonicum), which is a major causative agent of schistosomiasis. It is a severe disease globally affecting humans and animals. To test this hypothesis, we firstly conducted whole mount in situ hybridization analyses and found that the S. japonicum Vasa3 (SjVasa3) gene was expressed mainly in the reproductive organs...
January 5, 2018: Gene
https://www.readbyqxmd.com/read/28936657/whole-mount-rna-fish-on-ovules-and-developing-seeds
#10
Andrea Bleckmann, Thomas Dresselhaus
A key element to understand developmental and reproductive processes like germline development, double fertilization, and embryogenesis is the study of cell-specific gene expression patterns which is best analyzed by RNA in situ hybridization. Different visualization techniques have been established to mark either the region of mRNA production (using the classical chromogenic detection system) or the specific localization of mRNAs (using fluorescent labeled probes). In this chapter, we describe and compare whole mount RNA in situ hybridization techniques on ovules and young developing seeds from Arabidopsis thaliana using three different detection systems...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28891223/promoter-architecture-and-transcriptional-regulation-of-musculoskeletal-embryonic-nuclear-protein-1b-mustn1b-gene-in-zebrafish
#11
Paula Suarez-Bregua, Chien-Ju Chien, Manuel Megias, Shaojun Du, Josep Rotllant
BACKGROUND: Mustn1 is a specific musculoskeletal protein that plays a critical role in myogenesis and chondrogenesis in vertebrates. Whole-mount in situ hybridization revealed that mustn1b mRNAs are specifically expressed in skeletal and cardiac muscles in Zebrafish embryos. However, the precise function and the regulatory elements required for its muscle-specific expression are largely unknown. RESULTS: The purpose of this study was to explore and uncover the target genomic regions that regulate mustn1b gene expression by in vivo functional characterization of the mustn1b promoter...
September 11, 2017: Developmental Dynamics: An Official Publication of the American Association of Anatomists
https://www.readbyqxmd.com/read/28855284/whole-genome-sequencing-based-mapping-and-candidate-identification-of-mutations-from-fixed-zebrafish-tissue
#12
Nicholas E Sanchez, Breanne L Harty, Thomas O'Reilly-Pol, Sarah D Ackerman, Amy L Herbert, Melanie Holmgren, Stephen L Johnson, Ryan S Gray, Kelly R Monk
As forward genetic screens in zebrafish become more common, the number of mutants that cannot be identified by gross morphology or through transgenic approaches, such as many nervous system defects, has also increased. Screening for these difficult-to-visualize phenotypes demands techniques such as whole-mount in situ hybridization (WISH) or antibody staining, which require tissue fixation. To date, fixed tissue has not been amenable for generating libraries for whole genome sequencing (WGS). Here, we describe a method for using genomic DNA from fixed tissue and a bioinformatics suite for WGS-based mapping of zebrafish mutants...
October 5, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28828177/novel-bacteriocyte-associated-pleomorphic-symbiont-of-the-grain-pest-beetle-rhyzopertha-dominica-coleoptera-bostrichidae
#13
Genta Okude, Ryuichi Koga, Toshinari Hayashi, Yudai Nishide, Xian-Ying Meng, Naruo Nikoh, Akihiro Miyanoshita, Takema Fukatsu
BACKGROUND: The lesser grain borer Rhyzopertha dominica (Coleoptera: Bostrichidae) is a stored-product pest beetle. Early histological studies dating back to 1930s have reported that R. dominica and other bostrichid species possess a pair of oval symbiotic organs, called the bacteriomes, in which the cytoplasm is densely populated by pleomorphic symbiotic bacteria of peculiar rosette-like shape. However, the microbiological nature of the symbiont has remained elusive. RESULTS: Here we investigated the bacterial symbiont of R...
2017: Zoological Letters
https://www.readbyqxmd.com/read/28826993/expression-and-characterization-of-the-zebrafish-orthologue-of-the-human-folr1-gene-during-embryogenesis
#14
RoJenia N Jones, Stephanie A Erhard, Mark R Malham, Ayaz Y Gen, Kyle Sullivan, Kenneth W Olsen, Rodney M Dale
It has been well established that many types of rapidly dividing normal and diseased cells require an increased amount of folate for DNA replication and repair as well as cellular metabolism. Thus one of folate's cognate receptors, Folate Receptor 1 (FOLR1) is usually up-regulated in rapidly dividing cells, including many types of cancerous tumors. Because zebrafish have become a model organism for understanding conserved vertebrate cellular pathways and human disease, there has been an increased need to identify and elucidate orthologous zebrafish genes that are central to known human maladies...
November 2017: Gene Expression Patterns: GEP
https://www.readbyqxmd.com/read/28791673/comprehensive-analyses-of-hox-gene-expression-in-xenopus-laevis-embryos-and-adult-tissues
#15
Mariko Kondo, Takayoshi Yamamoto, Shuji Takahashi, Masanori Taira
From whole genome sequencing of an allotetraploid frog, Xenopus laevis, two homeologous sets (L and S) of four Hox clusters A through D (HoxA.L/S, HoxB.L/S, HoxC.L/S, and HoxD.L/S) and 13 paralogous groups (PGs) with 76 genes were identified, allowing us to carry out the first comprehensive analyses of hox gene expression in vertebrates. Expression of all hox genes during development and in adult tissues was analyzed by RNA-sequencing. The expression levels of most hox genes were similar between homeologs, but in some pairs, large differences were observed and several of these were confirmed by RT-PCR and whole mount in situ hybridization experiments...
August 9, 2017: Development, Growth & Differentiation
https://www.readbyqxmd.com/read/28765643/mir-202-5p-is-a-novel-germ-plasm-specific-microrna-in-zebrafish
#16
Jing Zhang, Wei Liu, Yilin Jin, Peng Jia, Kuntong Jia, Meisheng Yi
Gametogenesis is a complicated biological process by which sperm and egg are produced for genetic transmission between generations. In many animals, the germline is segregated from the somatic lineage in early embryonic development through the specification of primordial germ cells (PGCs), the precursors of gametes for reproduction and fertility. In some species, such as fruit fly and zebrafish, PGCs are determined by the maternally provided germ plasm which contains various RNAs and proteins. Here, we identified a germ plasm/PGC-specific microRNA miR-202-5p for the first time in zebrafish...
August 1, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28735975/molecular-characterization-and-expression-profiles-of-myosin-essential-light-chain-gene-in-the-pacific-oyster-crassostrea-gigas
#17
Hong Yu, Huijuan Li, Qi Li
In molluscs, muscle contraction is triggered by a direct binding of Ca(2+) to myosin. The myosin essential light chain (MELC) provides the Ca(2+) binding site and is of importance for motor function and regulation. In this study, the complete cDNA sequence of MELC gene of the Pacific oyster, Crassostrea gigas, was obtained, and the expression profiles were performed in different tissues and different embryo-larval development stages. The results showed that the full length of C. gigas MELC (CgMELC) cDNA was 659bp, containing a 5'-untranslated region of 73 bp, a 3'-untranslated region of 112bp...
November 2017: Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology
https://www.readbyqxmd.com/read/28725656/expression-of-vox-and-vent-mrnas-and-encoded-proteins-in-zebrafish-embryos
#18
Elena S Pshennikova, Maria B Tereshina, Anna S Voronina
In Danio rerio (zebrafish), members of the vent gene-family (vox/vega1, vent/vega2) are considered as ventralizing factors. We investigated not only the expression of their mRNAs by in situ hybridization at different stages of embryonic development, but also the spatial distribution of the encoded proteins by whole-mount immunostaining. We showed vox mRNA to be available in embryos since early cleavage and later on. Vent mRNA appeared after zygotic genome activation only. The vox and vent proteins were revealed at stage of eight blastomeres...
2017: Stem Cell Investigation
https://www.readbyqxmd.com/read/28724733/early-evolution-of-radial-glial-cells-in-bilateria
#19
Conrad Helm, Anett Karl, Patrick Beckers, Sabrina Kaul-Strehlow, Elke Ulbricht, Ioannis Kourtesis, Heidrun Kuhrt, Harald Hausen, Thomas Bartolomaeus, Andreas Reichenbach, Christoph Bleidorn
Bilaterians usually possess a central nervous system, composed of neurons and supportive cells called glial cells. Whereas neuronal cells are highly comparable in all these animals, glial cells apparently differ, and in deuterostomes, radial glial cells are found. These particular secretory glial cells may represent the archetype of all (macro) glial cells and have not been reported from protostomes so far. This has caused controversial discussions of whether glial cells represent a homologous bilaterian characteristic or whether they (and thus, centralized nervous systems) evolved convergently in the two main clades of bilaterians...
July 26, 2017: Proceedings. Biological Sciences
https://www.readbyqxmd.com/read/28716076/klf8-regulates-left-right-asymmetric-patterning-through-modulation-of-kupffer-s-vesicle-morphogenesis-and-spaw-expression
#20
Che-Yi Lin, Ming-Yuan Tsai, Yu-Hsiu Liu, Yu-Fen Lu, Yi-Chung Chen, Yun-Ren Lai, Hsin-Chi Liao, Huang-Wei Lien, Chung-Hsiang Yang, Chang-Jen Huang, Sheng-Ping L Hwang
BACKGROUND: Although vertebrates are bilaterally symmetric organisms, their internal organs are distributed asymmetrically along a left-right axis. Disruption of left-right axis asymmetric patterning often occurs in human genetic disorders. In zebrafish embryos, Kupffer's vesicle, like the mouse node, breaks symmetry by inducing asymmetric expression of the Nodal-related gene, spaw, in the left lateral plate mesoderm (LPM). Spaw then stimulates transcription of itself and downstream genes, including lft1, lft2, and pitx2, specifically in the left side of the diencephalon, heart and LPM...
July 17, 2017: Journal of Biomedical Science
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