RNA Interference and CRISPR Technologies Technical Advances and New Therapeutic Opportunities

This volume explores the uses of RNAi and CRISPR interferences as a general method for inhibiting gene expression, with focus on their biological functions, design, chemical modifications, delivery, and preclinical/clinical applications. In addition to relevant backgrounds, the chapters in this book discuss simple and accurate protocols dealing with the latest advances in RNAi and CRISPR applications and look at how these methods differ from one another. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

RNA and CRISPR Interferences: Past, Present and Future Perspectives.- Chemical Modifications in RNA Interference and CRISPR/Cas Genome Editing Reagents.- Preparation, Determination of Activity, and Biodistribution of Cholesterol-Containing Nuclease-Resistant siRNAs In Vivo.- Multifunctional Nanodelivery Platform for Maximizing Nucleic Acids Combination Therapy.- PAMAM Dendrimers as a Delivery System for Small Interfering RNA.- Delivery of Functional Small RNAs via Extracellular Vesicles In Vitro and In Vivo.- Optimized siRNA Delivery into Primary Immune Cells using Electroporation.- Synthesis and Evaluation of Caged siRNAs with Single cRGD Modification for Photoregulating RNA Interference.- Exploring 5’-Biotinylation of the Sense Strand to Improve siRNA Specificity and Potency.- In Vivo Delivery of Cassettes Encoding Anti-HBV Primary MicroRNAs using an Ancestral Adeno-Associated Viral Vector.- Generating DNA Expression Cassettes Encoding Multimeric Artificial MicroRNA Precursors.- MiR-302-Mediated Somatic Cell Reprogramming and Method for Generating Tumor-Free iPS Cells Using miR-302.- Urinary MicroRNAs as Emerging Class of Noninvasive Biomarkers.- Improving Dendritic Cell Cancer Vaccine Potency using RNA Interference.- Unleashing the Therapeutic Potential of Dendritic and T Cell Therapies using RNA Interference.- Harnessing the Antiviral-Type Responses Induced by Immunostimulatory siRNAs for Cancer Immunotherapy.- Cancer Immunotherapy: Targeting Tumor-Associated Macrophages by Gene Silencing.- Use of RNA Interference with TCR Transfer Enhance Safety and Efficiency.- CRISPR/Cas9 Guide RNA Design Rules for Predicting Activity.- CRISPR/Cas9 Genome Editing in Human Cell Lines with DONOR Vector Made by Gibson Assembly.- Genome Editing in Zebrafish using High-Fidelity Cas9 Nucleases: Choosing the Right Nuclease for the Task.- Next Generation of Adoptive T Cell Therapy using CRISPR/Cas9 Technology: Universal or Boosted?.- Engineering T-Cells using CRISPR/Cas9 for Cancer Therapy.- CRISPR/Cas9 Mediated Genome Engineering of Primary Human B Cells.- Gene Editing in B-Lymphoma Cell Lines using CRISPR/Cas9 Technology.- Gene Knockout in Hematopoietic Stem and Progenitor Cells followed by Granulocytic Differentiation.- CRISPR/Cas9 Genome Editing of Human Induced Pluripotent Stem Cells followed by Granulocytic Differentiation.