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Experiment Tags: Recombinant Virus, Vaccinia Virus, VACV (VACV), CRISPR/Cas9 Selection, Virotherapy, Vaccine Development
Experiment Summary: In this protocol, we describe protocols for the generation and enrichment of recombinant vaccinia viruses using targeted Cas9 as a selection tool. This novel use of Cas9 is a simple addition to current homologous recombination-based methods that are widespread in the field, facilitating implementation in laboratories already working with poxviruses.
Experiment Tags: Subcellular Fractionation, RT-qPCR, Circular RNA, circRNA, Cytoplasmic circRNA, Nuclear circRNA, Low-Abundance circRNA
Experiment Summary: In this protocol, we present a protocol for the subcellular fractionation of cells to detect circRNA via RT-qPCR with divergent primers. Moreover, detailed steps for the generation of specific circRNAs-enriched cDNA included in this protocol will enhance the amplification and detection of low-abundance circRNAs.
Experiment Tags: dsRNA Synthesis, In Vitro Preparation
Experiment Summary: In this protocol, we describe a simple and widely used in vitro transcription reaction for the preparation of double-stranded RNA using PCR templates. The dsRNA prepared by this method can be used to induce RNA interference in certain cells or tissues.
Experiment Tags: CRISPR/Cas9 (CRISPR/ Cas9), Adeno-Associated Viral Vector, Trans-Zona Pellucida, Intra-Embryo Genome Editing, Ribonucleoprotein Electroporation, Large Fragment Knock-In
Experiment Summary: In this protocol, we demonstrate a detailed protocol for AAV-mediated intra-embryo knock-in in laboratory mice. This technique does not require either micromanipulation or germline competent pluripotent stem cells.
Experiment Tags: Biological Techniques, Recombinant, Lentivirus, Transduction
Experiment Summary: In this protocol, we describe the transduction of cultured cells with recombinant lentivirus expressing fluorescent markers.
Experiment Tags: Nucleoside-Modified mRNA, Lipid Nanoparticle, Liver Regeneration, Protein Expression, Retro-Orbital Injection
Experiment Summary: In this protocol, we describe in detail the necessary steps to deliver mRNA-LNP to the murine liver and, following intravenous injection of eGFP mRNA-LNP, verify transfection efficiency using flow cytometry and liver cell specificity using immunofluorescence analyses
Experiment Tags: Rubella Virus, Lipid, Sphingomyelin, Virus Entry, Sphingomyelinase
Experiment Summary: This protocol evaluates in detail the effect of SMase treatment on RuV infectivity in RK13 and Vero cells.
Experiment Tags: CRISPR-Cas9, Off-Target Activity, Chromosomal Translocation, LAM-HTGTS, iHTGTS
Experiment Summary: In this article, we described an improved HTGTS (iHTGTS) method, in which size-selection beads were used to enhance reaction efficiency and a new primer system was designed to be compatible with Illumina Hiseq sequencing. iHTGTS is a powerful method for comprehensively assessing Cas9 off-target effect.
Experiment Tags: Diagnosis, RNA Virus, Replicative Intermediate, dsRNA Isolation, dsRNA-Binding Protein
Experiment Summary: In this protocol, we describe a method for dsRNA isolation. Using this method, we can readily isolate viral dsRNA from a small amount of plant material, and can process numerous samples simultaneously.
Experiment Tags: Single Guide RNA Library, Library Design and Construction
Experiment Summary: This protocol describes how to generate a single guide RNA (sgRNA) library for use in genetic screening.
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