Genome Editing Techniques

This video explains the concept of CRISPR-mediated cytosine base editors for inducing targeted nucleotide substitution.

In this video, we demonstrate all-in-one CRISPR-Cas9 based genome editing in cultured cells where Cas9 and sgRNA are provided as a single plasmid construct to the cells. The CRISPR-Cas9 system and desired gene to be inserted was introduced in cells through electroporation technique to facilitate successful gene editing.

This video demonstrates a technique of zinc-finger nuclease-based genome editing in human pluripotent stem cells (hPSCs). As hPSCs can potentially be differentiated into various cell types, this technology is helpful for in vitro pathological and pharmacological studies in a patient-specific manner.

This video describes a precise genome editing technique in human induced pluripotent stem cells, or hiPSCs, using TALENs to create double-stranded breaks at a targeted locus, inducing homology-directed repair for fluorescence protein gene integration.

In this video, we demonstrate phiC31-mediated site-specific transgene integration in the Anopheles embryo via microinjection. This method helps introduce desired genes into a host system for research and industrial applications.

In this video, we demonstrate generation of transgenic Arabidopsis plants expressing stable transgene through Agrobacterium-mediated binary bacterial artificial chromosome-based gene transformation.

In this video, we demonstrate a procedure to generate a stable zebrafish transgenic line constitutively expressing a fluorescent voltage indicator protein following the co-injection of the Tol2 transposon system comprising the transposase mRNA and Tol2 transposon construct encoding the fluorescent voltage indicator.

This video demonstrates the procedure to generate motor neurons by transfection of human iPSCs with PiggyBac transposon system by the ectopic expression of lineage-specific transcription factors.

This video demonstrates a CRISPR interference-based gene silencing in pathogenic Leptospira. Targeted repression of gene expression using a CRISPR/Cas system that blocks mRNA formation helps identify candidate genes for pathological and pharmacological studies.

This video describes a procedure for target gene knockout using a tamoxifen-inducible Cre-recombinase system following intraperitoneal injection of tamoxifen in a mouse model.

This video describes a gene knockout technique using a CRISPR-concatemer to simultaneously knock out multiple genes in cultured mouse intestinal organoid cells. This method is used to knock out a diseased gene and to elucidate the function of a gene and its paralogues.

In this video, we demonstrate a method to perform transduction of shRNA lentiviral vectors to obtain stable knockdown cell lines in immortalized hematopoietic stem and progenitor cells (iHSPCs).

This video demonstrates the tetracycline, or Tet-On, gene expression system in a mouse model. In the presence of doxycycline, a tetracycline derivative, the Tet-On system consisting of reverse tetracycline-controlled transactivator protein becomes activated and expresses the gene of interest.

In this video, we describe a procedure to perform reverse transfection of siRNA into adipocytes to modulate the expression of a target protein.

This video describes the method of regulating gene expression in cultured cells by transfection with a plasmid containing a gene of interest and a regulatory plasmid. The expression of this plasmid can be controlled by tetracycline or its synthetic analogs, making this system a useful model to study regulatable gene expression.

This video describes a gene knockdown technique using translational blocking morpholinos microinjected in the one-cell stage of the cavefish. Morpholinos are DNA oligonucleotides that bind to the complementary sequence of target mRNA and block its translation to a protein. Consequently, morpholinos are used as molecular tools to stop or reduce a gene’s expression transiently.