Molecular Biology

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    Protocols in Current Issue
    In situ Hybridization of Plant-parasitic Nematode Globodera pallida Juveniles to Detect Gene Expression
    In this study, we describe a standard whole mount in situ hybridization method which is used to determine the spatial-temporal expression pattern of genes from Globodera spp. Unlike more invasive radioactive labeling approaches, ...
    A PhoA-STII Based Method for Efficient Extracellular Secretion and Purification of Fab from Escherichia coli
    Authors:  Ziyan Wang, Yang Gao, Manyu Luo, Cedric Cagliero, Hua Jiang, Yueqing Xie, Jianwei Zhu and Huili Lu, date: 09/20/2019, view: 24, Q&A: 0
    In comparison with full-length IgGs, antigen binding fragments (Fabs) are smaller in size and do not require the complexed post-translational modification. Therefore, Fab can be cost-effectively produced using an Escherichia coli (E. coli ...
    Using indCAPS to Detect CRISPR/Cas9 Induced Mutations
    Authors:  Charles Hodgens, Zachary L. Nimchuk and Joseph J. Kieber, date: 09/20/2019, view: 27, Q&A: 0
    Cleaved amplified polymorphic sequences (CAPS) assays are useful tools for detecting small mutations such as single nucleotide polymorphisms (SNPs) or insertion/deletions (indels) present in an amplified DNA fragment. A mutation that disrupts or ...
    In situ Hybridization of Plant-parasitic Nematode Globodera pallida Juveniles to Detect Gene Expression
    [Abstract] In this study, we describe a standard whole mount in situ hybridization method which is used to determine the spatial-temporal expression pattern of genes from Globodera spp. Unlike more invasive radioactive labeling approaches, this technique is based on a safe, highly specific enzyme-linked immunoassay where a Digoxigenin ...
    A PhoA-STII Based Method for Efficient Extracellular Secretion and Purification of Fab from Escherichia coli
    Authors:  Ziyan Wang, Yang Gao, Manyu Luo, Cedric Cagliero, Hua Jiang, Yueqing Xie, Jianwei Zhu and Huili Lu, date: 09/20/2019, view: 24, Q&A: 0
    [Abstract] In comparison with full-length IgGs, antigen binding fragments (Fabs) are smaller in size and do not require the complexed post-translational modification. Therefore, Fab can be cost-effectively produced using an Escherichia coli (E. coli) expression system. However, the disulfide-bonds containing exogenous protein, including ...
    Using indCAPS to Detect CRISPR/Cas9 Induced Mutations
    Authors:  Charles Hodgens, Zachary L. Nimchuk and Joseph J. Kieber, date: 09/20/2019, view: 27, Q&A: 0
    [Abstract] Cleaved amplified polymorphic sequences (CAPS) assays are useful tools for detecting small mutations such as single nucleotide polymorphisms (SNPs) or insertion/deletions (indels) present in an amplified DNA fragment. A mutation that disrupts or creates a restriction site will prevent cleavage by a restriction enzyme, allowing discrimination of ...
    Yeast Single-cell RNA-seq, Cell by Cell and Step by Step
    [Abstract] Single-cell RNA-seq (scRNA-seq) has become an established method for uncovering the intrinsic complexity within populations. Even within seemingly homogenous populations of isogenic yeast cells, there is a high degree of heterogeneity that originates from a compact and pervasively transcribed genome. Research with microorganisms such as yeast ...
    A Refined Protocol for Identifying Citrulline-specific Monoclonal Antibodies from Single Human B Cells from Rheumatoid Arthritis Patient Material
    [Abstract] We describe here a detailed, refined protocol for the generation of citrulline-specific monoclonal antibodies from single human B cells from rheumatoid arthritis (RA) patients. This protocol provides a detailed guide of the procedure starting from single B cells of your choice and followed by amplification of the variable region of immunoglobulin ...
    PEG-mediated, Stable, Nuclear and Chloroplast Transformation of Cyanidioschizon merolae
    Authors:  Maksymilian Zienkiewicz, Tomasz Krupnik, Anna Drożak and Kinga Kania, date: 09/05/2019, view: 150, Q&A: 0
    [Abstract] The ability to achieve nuclear or chloroplast transformation in plants has been a long standing goal, especially in microalgae research. Over past years there has been only little success, but transient and stable nuclear transformation has been achieved in multiple species. Our newly developed method allows for relatively simple transformation of ...
    Development of an Efficient Transformation System for Halotolerant Yeast Debaryomyces hansenii CBS767
    Authors:  Anu P. Minhas and Dipanwita Biswas, date: 09/05/2019, view: 154, Q&A: 0
    [Abstract] Debaryomyces hansenii is one of the most osmotolerant and halotolerant yeasts. Further, its association with traditional cheese and meat products imparting special flavors to these products project this yeast with enormous biotechnological potential in the agrofood sector. However, lack of an efficient transformation system in D. hansenii ...
    A Novel Dual Lentiviral CRISPR-based Transcriptional Activation System for Gene Expression Regulation in Neurons
    Authors:  Katherine E. Savell, Faraz A. Sultan and Jeremy J. Day, date: 09/05/2019, view: 563, Q&A: 0
    [Abstract] Robust and efficient gene expression control enables the study of a gene’s function in the central nervous system. Advances in CRISPR-based technology provide new avenues not only for gene editing, but for complex transcriptional control. Here, we describe a protocol to generate high-titer lentiviruses with neuron-optimized CRISPR-activation ...
    Application of a Modified Smart-seq2 Sample Preparation Protocol for Rare Cell Full-length Single-cell mRNA Sequencing to Mouse Oocytes
    Authors:  Rebecca S. Treger, Scott D. Pope, Xiaojun Xing and Akiko Iwasaki, date: 08/20/2019, view: 281, Q&A: 0
    [Abstract] Endogenous retroviruses (ERV) are transposable retroelements that form ~10% of the murine genome and whose family members are differentially expressed throughout embryogenesis. However, precise regulation of ERV in germ cells remains unclear. To investigate ERV expression in oocytes, we adapted a single-cell mRNA-sequencing library preparation ...
    Construction of Viral Vectors for Cell Type-specific CRISPR Gene Editing in the Adult Mouse Brain
    Authors:  Hiroshi Yamaguchi and Luis de Lecea, date: 08/20/2019, view: 452, Q&A: 0
    [Abstract] Recently developed gene editing technologies based on engineered CRISPR/Cas9 systems enables researchers to disrupt genes in a cell type-specific manner in the adult mouse brain. Using these technologies, we recently showed that the dopamine beta-hydroxylase gene in Locus Coeruleus (LC) norepinephrine neurons plays a vital role in the maintenance ...



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