Developmental Biology

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    Protocols in Current Issue
    Direct-TRI: High-throughput RNA-extracting Method for all Stages of Zebrafish Development
    Authors:  Kota Ujibe, Kanako Nishimura, Makoto Kashima and Hiromi Hirata, date: 09/05/2021, view: 400, Q&A: 0

    Recent popularization of next-generation sequencing enables conducting easy transcriptome analysis. Nevertheless, substantial RNA isolation work prior to RNA sequencing, as well as the high cost involved, still makes the routine use of large-scale

    ...
    Isolation of Myofibres and Culture of Muscle Stem Cells from Adult Zebrafish
    Authors:  Massimo Ganassi, Peter S. Zammit and Simon M. Hughes, date: 09/05/2021, view: 1034, Q&A: 0

    Skeletal muscles generate force throughout life and require maintenance and repair to ensure efficiency. The population of resident muscle stem cells (MuSCs), termed satellite cells, dwells beneath the basal lamina of adult myofibres and contributes

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    In situ Hybridization of miRNAs in Human Embryonic Kidney and Human Pluripotent Stem Cell-derived Kidney Organoids
    Authors:  Filipa M. Lopes, Susan J. Kimber and Ioannis Bantounas, date: 09/05/2021, view: 341, Q&A: 0

    MicroRNAs are small RNAs that negatively regulate gene expression and play an important role in fine-tuning molecular pathways during development. There is increasing interest in studying their function in the kidney, but the majority of studies to

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    In situ Hybridization of miRNAs in Human Embryonic Kidney and Human Pluripotent Stem Cell-derived Kidney Organoids
    Authors:  Filipa M. Lopes, Susan J. Kimber and Ioannis Bantounas, date: 09/05/2021, view: 341, Q&A: 0
    [Abstract]

    MicroRNAs are small RNAs that negatively regulate gene expression and play an important role in fine-tuning molecular pathways during development. There is increasing interest in studying their function in the kidney, but the majority of studies to date use kidney cell lines and assess the total amounts of miRNAs of interest either by qPCR or by

    ...
    Isolation of Myofibres and Culture of Muscle Stem Cells from Adult Zebrafish
    Authors:  Massimo Ganassi, Peter S. Zammit and Simon M. Hughes, date: 09/05/2021, view: 1034, Q&A: 0
    [Abstract]

    Skeletal muscles generate force throughout life and require maintenance and repair to ensure efficiency. The population of resident muscle stem cells (MuSCs), termed satellite cells, dwells beneath the basal lamina of adult myofibres and contributes to both muscle growth and regeneration. Upon exposure to activating signals, MuSCs proliferate to

    ...
    Direct-TRI: High-throughput RNA-extracting Method for all Stages of Zebrafish Development
    Authors:  Kota Ujibe, Kanako Nishimura, Makoto Kashima and Hiromi Hirata, date: 09/05/2021, view: 400, Q&A: 0
    [Abstract]

    Recent popularization of next-generation sequencing enables conducting easy transcriptome analysis. Nevertheless, substantial RNA isolation work prior to RNA sequencing, as well as the high cost involved, still makes the routine use of large-scale transcriptome analysis difficult. For example, conventional phenol-chloroform RNA extraction cannot

    ...
    Preparation of Drosophila Larval Blood Cells for Single-cell RNA Sequencing
    Authors:  Sudhir Gopal Tattikota and Norbert Perrimon, date: 08/20/2021, view: 464, Q&A: 0
    [Abstract]

    Recent advances in single-cell RNA-sequencing (scRNA-seq) technologies provide unprecedented opportunities to identify new cell types and characterize cell states. One of the most important requirements for performing scRNA-seq is to obtain high-quality single cells in suspension. Recently, we used this approach to characterize Drosophila blood

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    Measuring DNA Damage Using the Alkaline Comet Assay in Cultured Cells
    Authors:  Elena Clementi, Zuzana Garajova and Enni Markkanen, date: 08/20/2021, view: 888, Q&A: 0
    [Abstract]

    Maintenance of DNA integrity is of pivotal importance for cells to circumvent detrimental processes that can ultimately lead to the development of various diseases. In the face of a plethora of endogenous and exogenous DNA-damaging agents, cells have evolved a variety of DNA repair mechanisms that are responsible for safeguarding genetic

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    A Genetically Engineered Mouse Model of Venous Anomaly and Retinal Angioma-like Vascular Malformation
    Authors:  Xudong Cao, Beibei Xu, Xiao Li, Taotao Li and Yulong He, date: 08/05/2021, view: 744, Q&A: 0
    [Abstract]

    Characterization of key regulators in vein development will advance our understanding of mechanisms underlying venous anomalies and provide therapeutic targets for the treatment of vascular malformations. Here, we provide a detailed protocol for the generation of genetically engineered mouse models targeting the Tek gene for the analysis of vein

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    Fe-NTA Microcolumn Purification of Phosphopeptides from Immunoprecipitation (IP) Eluates for Mass Spectrometry Analysis
    Authors:  Ethan J. Sanford and Marcus B. Smolka, date: 08/05/2021, view: 487, Q&A: 0
    [Abstract]

    Protein phosphorylation is a nearly universal signaling mechanism. To date, a number of proteomics tools have been developed to analyze phosphorylation. Phosphoproteome-wide analyses using whole cell extracts suffer from incomplete coverage, often missing phosphorylation events from low-abundance proteins. In order to increase coverage of

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    Mouse Periosteal Cell Culture, in vitro Differentiation, and in vivo Transplantation in Tibial Fractures
    [Abstract]

    The periosteum covering the outer surface of bone contains skeletal stem/progenitor cells that can efficiently form cartilage and bone during bone repair. Several methods have been described to isolate periosteal cells based on bone scraping and/or enzymatic digestion. Here, we describe an explant culture method to isolate periosteum-derived

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    Isolation and in vitro Culture of Mouse Oocytes
    Authors:  Jessica Greaney, Goutham Narayanan Subramanian, Yunan Ye and Hayden Homer, date: 08/05/2021, view: 850, Q&A: 0
    [Abstract]

    Females are endowed at birth with a fixed reserve of oocytes, which declines both in quantity and quality with advancing age. Understanding the molecular mechanisms regulating oocyte quality is crucial for improving the chances of pregnancy success in fertility clinics. In vitro culture systems enable researchers to analyse important molecular and

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    GeneWeld: Efficient Targeted Integration Directed by Short Homology in Zebrafish
    [Abstract]

    Efficient precision genome engineering requires high frequency and specificity of integration at the genomic target site. Multiple design strategies for zebrafish gene targeting have previously been reported with widely varying frequencies for germline recovery of integration alleles. The GeneWeld protocol and pGTag (plasmids for Gene Tagging)

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