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    CRISPR - Cas System

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    meghna
    The document summarizes the discovery and mechanism of the CRISPR-Cas genome editing system. It describes how CRISPR sequences were initially discovered in bacteria and later found to provide adaptive immunity. The system works by integrating short DNA sequences from viruses/plasmids between repeat sequences. This allows the Cas proteins to recognize and cut invading DNA for degradation. The document outlines applications of CRISPR in gene knockout, knock-in, and interference in eukaryotic systems including potential medical uses.

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    CRISPR - Cas System

    Uploaded by

    meghna
    212 views13 pages
    The document summarizes the discovery and mechanism of the CRISPR-Cas genome editing system. It describes how CRISPR sequences were initially discovered in bacteria and later found to provide adaptive immunity. The system works by integrating short DNA sequences from viruses/plasmids between repeat sequences. This allows the Cas proteins to recognize and cut invading DNA for degradation. The document outlines applications of CRISPR in gene knockout, knock-in, and interference in eukaryotic systems including potential medical uses.

    Original Description:

    the new method of genome editing.

    Original Title

    CRISPR- Cas System

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    The document summarizes the discovery and mechanism of the CRISPR-Cas genome editing system. It describes how CRISPR sequences were initially discovered in bacteria and later found to provide adaptive immunity. The system works by integrating short DNA sequences from viruses/plasmids between repeat sequences. This allows the Cas proteins to recognize and cut invading DNA for degradation. The document outlines applications of CRISPR in gene knockout, knock-in, and interference in eukaryotic systems including potential medical uses.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    212 views13 pages

    CRISPR - Cas System

    Uploaded by

    meghna
    The document summarizes the discovery and mechanism of the CRISPR-Cas genome editing system. It describes how CRISPR sequences were initially discovered in bacteria and later found to provide adaptive immunity. The system works by integrating short DNA sequences from viruses/plasmids between repeat sequences. This allows the Cas proteins to recognize and cut invading DNA for degradation. The document outlines applications of CRISPR in gene knockout, knock-in, and interference in eukaryotic systems including potential medical uses.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    of 13

    CRISPR- CAS

    SYSTEM

    A powerful new tool for GENOME EDITIN


    Submitted to:
    Dr. T.C.Bhalla

    Submitted by:
    Meghna Garg
    R.NO: 3401

    DISCOVERY OF CRISPR-CaS SYSTEM

    In 1987, the first description of a CRISPR locus


    was published by a lab which discovered a
    group of 29-nucleotide repeats divided by nonrepetitive short sequences in E. coli, which are
    now known as spacers (Ishino et al., 1987).
    Mojica et al. (2000) later determined that
    similar sequences of short regularly spaced
    repeats (SRSRs) are present in many species of
    bacteria. In 2002, Jansen et al. further defined
    these SRSRs as a group of repeats that is
    preceded by both aleader sequence and an
    AT-rich region.

    The authors also found a conserved family of genes that


    generally exist near the repeats. This family consisted of
    genes encoding proteins like DNA helicases (cas3) and
    exonucleases (cas4), which implied that Cas genes
    likely have function in DNA metabolism. They renamed
    the SRSRs as CRISPRs and the genes found near them
    as CRISPR-associated genes (Cas) (Jansen et al, 2002).
    Bolotin et al (2005) sequenced the CRISPR spacers of 24
    strains of Streptococcus thermophilus and
    Streptococcus vestibularis and found that the
    sequences were homologous with extra-chromosomal
    elements. The authors theorized that CRISPR spacers
    are the heritable remnants of infection from exogenous
    DNA.

    INTEGRATION OF
    SPACERS

    MECHANISM OF
    FUNCTIONING

    FINAL BINDING

    APPLICATIONS IN
    EUKARYOTES

    Recent work has proven that the CRISPR/Cas9


    system can be utilized for gene editing in a
    plethora of systems including yeast, rice, zebra
    fish, mouse, and even humans.(Mali et al., 2013)
    Based on the type II CRISPR/Cas9 mechanism,
    researchers have engineered a RNA chimera of
    tracrRNA-crRNA called single guide RNA (sgRNA)
    which is able to cause sequence-specific binding
    to dsDNA (Cong et al, 2013). Cas9 nuclease is
    capable of interacting with this engineered
    sgRNA in order to specifically cleave dsDNA in
    regions complementary to the introduced sgRNA.

    1.GENE KNOCK OUT AND 2. KNOCK IN

    APPLICATIONS OF GENE KNOCK OUT

    GENE KNOCK OUT SYSTEM: CRISPR could be used


    to delete harmful disease causing mutations in the
    human genome, especially in IVF embryos.
    Alternatively, the system could be used to
    knockout specific genes in model organisms in
    order to study diseases.
    Multiple genes could be knocked out at the same
    time.
    Gene knockout system has been used
    effectively for knockout of AIDS virus. HOW??
    ANSWER- target the LTRs instead of CCR5 proteins.

    APPLICATIONS OF GENE KNOCK- IN

    GENE KNOCK IN SYSTEM: Many genetic diseases


    could be cured by modifying genomic sequences of
    pluripotent stem cells of patients to express wildtype copies of the disease causing genes. This
    would allow for autologous stem cell therapies
    which reduces the risk of graft-host disease.
    Schwank et al, studied the use of CRISPR
    gene knock-in as a therapy for Cystic Fibrosis.
    They succeeded in correcting mutant CFTR DeltaF508 alleles (alleles with a mutation that causes
    cystic fibrosis) using the CRISPR/Cas9 mediated
    homologous recombination in intestinal stem cells.

    CRISPR INTERFERENCE

    In 2013, Qi et al. created a catalytically dead Cas9


    (dCas9), lacking endonuclease activity, in order to
    function in gene silencing as opposed to gene editing
    through DSBs. This method, called CRISPR interference
    (CRISPRi) halts mRNA synthesis by blocking RNA
    polymerase at the promoter region of the DNA.
    CRISPRi has the ability to silence multiple genes at the
    same time.
    The authors research determined that dCas9 could be
    used to positively or negatively regulate the
    expression of particular target genes that influence cell
    differentiation. CRISPRi could therefore be useful to
    investigate stem cell differentiation pathways
    (Kearns et al,2014).

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