Molecular Biology


    Protocols in Current Issue
    Ectopic Gene Expression in Macrophages Using in vitro Transcribed mRNA
    Authors:  Pallavi Chandra and Jennifer A. Philips, date: 05/20/2018, view: 4159, Q&A: 0
    [Abstract] Macrophages are immune cells that contribute to host defense through various mechanisms including phagocytosis and antigen presentation. Their antimicrobial capacity is subverted by clinically important intracellular pathogens such as Mycobacterium tuberculosis. The study of host-pathogen interactions using these cells is therefore of ...
    Multiplex Gene Editing via CRISPR/Cas9 System in Sheep
    Authors:  Yiyuan Niu, Yi Ding, Xiaolong Wang and Yulin Chen, date: 07/05/2017, view: 7268, Q&A: 0
    [Abstract] Sheep is a major large animal model for studying development and disease in biomedical research. We utilized CRISPR/Cas9 system successfully to modify multiple genes in sheep. Here we provide a detailed protocol for one-cell-stage embryo manipulation by co-injecting Cas9 mRNA and RNA guides targeting three genes (MSTN, ASIP, and ...
    A Protocol for Production of Mutant Mice Using Chemically Synthesized crRNA/tracrRNA with Cas9 Nickase and FokI-dCas9
    Authors:  Satoshi Hara, Miho Terao and Shuji Takada, date: 06/05/2017, view: 7326, Q&A: 0
    [Abstract] The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system is the most widely used genome editing tool. A common CRISPR/Cas9 system consists of two components: a single-guide RNA (sgRNA) and Cas9. Both components are required for the introduction of a double-strand break (DSB) at a specific ...
    Creating a RAW264.7 CRISPR-Cas9 Genome Wide Library
    Authors:  Brooke A Napier and Denise M Monack, date: 05/20/2017, view: 8874, Q&A: 0
    [Abstract] The bacterial clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 genome editing tools are used in mammalian cells to knock-out specific genes of interest to elucidate gene function. The CRISPR-Cas9 system requires that the mammalian cell expresses Cas9 endonuclease, guide RNA (gRNA) to lead the endonuclease to the gene of ...
    Ca2+ Measurements in Mammalian Cells with Aequorin-based Probes
    Authors:  Anna Tosatto, Rosario Rizzuto and Cristina Mammucari, date: 03/05/2017, view: 5593, Q&A: 0
    [Abstract] Aequorin is a Ca2+ sensitive photoprotein suitable to measure intracellular Ca2+ transients in mammalian cells. Thanks to recombinant cDNAs expression, aequorin can be specifically targeted to various subcellular compartments, thus allowing an accurate measurement of Ca2+ uptake and release of different ...
    Synthetic Lethality Screens Using RNAi in Combination with CRISPR-based Knockout in Drosophila Cells
    Authors:  Benjamin E. Housden, Hilary E. Nicholson and Norbert Perrimon, date: 02/05/2017, view: 7799, Q&A: 0
    [Abstract] A synthetic lethal interaction is a type of genetic interaction where the disruption of either of two genes individually has little effect but their combined disruption is lethal. Knowledge of synthetic lethal interactions can allow for elucidation of network structure and identification of candidate drug targets for human diseases such as cancer. ...
    Dharmacon siRNA Transfection of HeLa Cells
    Author:  Yanling Chen, date: 02/20/2012, view: 21298, Q&A: 0
    [Abstract] Small Interfering RNA (siRNA) is a class of double-stranded RNAs of 20-25 nucleotides that play important roles in many biological processes (Hamilton and Baulcombe, 1999). siRNAs act by “neutralizing” the mRNA of the target protein, facilitating degradation of the mRNA and hence altering the biological effect of the protein (reviewed in Hannon ...

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