Molecular Biology

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    Protocols in Current Issue
    A Novel Method to Construct Binary CRISPR Vectors for Plant Transformation by Single Round of PCR Amplification
    Authors:  Kang Li, Yuhui Wang and Chuanying Fang, date: 04/05/2021, view: 975, Q&A: 0
    [Abstract]

    CRISPR/Cas9 is an established and flexible tool for genome editing. However, most methods used to generate expression clones for the CRISPR/Cas9 are time-consuming. Hence, we have developed a one-step protocol to introduce sgRNA expression cassette(s) directly into binary vectors (Liu et al., 2020). In this approach, we have optimized the

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    Histochemical Staining of Suberin in Plant Roots
    Authors:  Peter Marhavý and Shahid Siddique, date: 02/05/2021, view: 1258, Q&A: 0
    [Abstract]

    Histological stains are useful tools for characterizing cell shape, arrangement and the material they are made from. Stains can be used individually or simultaneously to mark different cell structures or polymers within the same cells, and to visualize them in different colors. Histological stains can be combined with genetically-encoded

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    Generation and in Planta Functional Analysis of Potato Virus Y mutants
    [Abstract] Potato virus Y (PVY), the type member of the genus Potyvirus (family Potyviridae), is the most widespread virus affecting potato and is included in the top five most economically detrimental plant viruses. Recently, the structure of the PVY virion has been determined by cryo-electron microscopy, which has opened the doors to ...
    Assembly of Genetic Circuits with the Mammalian ToolKit
    Authors:  João P. Fonseca, Alain R Bonny, Jason Town and Hana El-Samad, date: 03/05/2020, view: 2477, Q&A: 0
    [Abstract] The ability to rapidly assemble and prototype cellular circuits is vital for biological research and its applications in biotechnology and medicine. The Mammalian ToolKit (MTK) is a Golden Gate-based cloning toolkit for fast, reproducible and versatile assembly of large DNA vectors and their implementation in mammalian models. The MTK consists of ...
    High-throughput Site-directed Scanning Mutagenesis Using a Two-fragment PCR Approach
    Authors:  Franziska M. Heydenreich, Tamara Miljuš, Dalibor Milić and Dmitry B. Veprintsev, date: 01/05/2020, view: 2941, Q&A: 0
    [Abstract] Site-directed scanning mutagenesis is a useful tool applied in studying protein function and designing proteins with new properties, such as increased stability or enzymatic activity. Creating a systematic library of hundreds of site-directed mutants is still a demanding and expensive task. The established protocols for making such libraries ...
    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 ...
    Multiple Modification of Chromosomal Loci Using URA5.3 Selection Marker in the Unicellular Red Alga Cyanidioschyzon merolae
    Authors:  Tokiaki Takemura, Sousuke Imamura, Yuki Kobayashi and Kan Tanaka, date: 04/05/2019, view: 3414, Q&A: 0
    [Abstract] The unicellular red alga Cyanidioschyzon merolae has been used as a eukaryotic photosynthetic model for various basic and applied studies. Although the nuclear genome of C. merolae can be modified by homologous recombination with exogenously introduced DNA, it has been difficult to modify multiple chromosome loci within the same ...
    Measuring Secretion of Capsidiol in Leaf Tissues of Nicotiana benthamiana
    Authors:  Teruhiko Kuroyanagi, Maurizio Camagna and Daigo Takemoto, date: 08/05/2018, view: 3434, Q&A: 0
    [Abstract] Plant species produce a wide variety of antimicrobial metabolites to protect themselves against potential pathogens in natural environments. Phytoalexins are low molecular weight compounds produced by plants in response to attempted attacks of pathogens. Accumulation of phytoalexins in attacked plant tissues can inhibit the growth of penetrating ...
    Precision Tagging: A Novel Seamless Protein Tagging by Combinational Use of Type II and Type IIS Restriction Endonucleases
    Authors:  Zhen Xu, Yan-Ning Rui, John P. Hagan and Dong H. Kim, date: 02/05/2018, view: 6223, Q&A: 0
    [Abstract] Protein tagging is a powerful tool for performing comprehensive analyses of the biological functions of a protein of interest owing to the existence of a wide variety of tags. It becomes indispensable in some cases, such as in tracking protein dynamics in a live cell or adding a peptide epitope due to the lack of optimal antibodies. However, ...
    Design and Direct Assembly of Synthesized Uracil-containing Non-clonal DNA Fragments into Vectors by USERTM Cloning
    [Abstract] This protocol describes how to order and directly assemble uracil-containing non-clonal DNA fragments by uracil excision based cloning (USER cloning). The protocol was generated with the goal of making synthesized non-clonal DNA fragments directly compatible with USERTM cloning. The protocol is highly efficient and would be compatible ...



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