Genetic Engineering and Gene Knockout
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Questions and Answers

What is the primary purpose of adding adaptors during the RNA sequencing process?

  • To capture fragments containing exons (correct)
  • To repair the ends of the RNA
  • To add poly A tails to the RNA
  • To enable PCR amplification

    • What is the function of the kanMX gene in targeted mutagenesis?

  • It catalyzes the PCR reaction
  • It provides a selective marker for the knockout construct (correct)
  • It converts ganciclovir into a toxic nucleotide
  • It searches for homology in the genome

    • What is the result of targeted mutagenesis by homologous recombination in yeast?

  • A knockout of a specific gene in a mammalian system
  • A transgenic organism with an introduced gene
  • A gene duplication event
  • A null mutant with a knockout of specific genes (correct)

    • What is the purpose of ES cells in targeted mutagenesis in mouse?

    <p>To facilitate homologous recombination</p> Signup and view all the answers

    What is the estimated size of the final data generated from Whole-Genome-Sequencing?

    <p>~10 GB</p> Signup and view all the answers

    What is the main function of the CRISPR-Cas9 system in bacteria?

    <p>To recognize and digest foreign DNA</p> Signup and view all the answers

    What is the role of PAM (protospacer adjacent motifs) in the CRISPR-Cas9 system?

    <p>To guide crRNA and tracrRNA to the gene of interest</p> Signup and view all the answers

    What is the result of using RNA interference (RNAi) in gene regulation?

    <p>The RNA expressed by the gene of interest is only degraded</p> Signup and view all the answers

    What is the difference between NHEJ and HDR in the CRISPR-Cas9 system?

    <p>NHEJ results in small insertions or deletions, while HDR results in precise changes</p> Signup and view all the answers

    What is an advantage of using the CRISPR-Cas9 system for genome editing?

    <p>It can edit multiple genes in one step</p> Signup and view all the answers

    What is the primary function of restriction enzymes like EcoRI in bacteria?

    <p>To defend against invading viruses</p> Signup and view all the answers

    What is the purpose of genetic complementation analysis in yeast?

    <p>To identify recessive mutations in the same or different genes</p> Signup and view all the answers

    What is the result of combining a DNA fragment with a vector in DNA cloning?

    <p>Recombinant DNA</p> Signup and view all the answers

    What is the purpose of using a plasmid as a vector in DNA cloning?

    <p>To provide sequences for maintaining the vector in the host</p> Signup and view all the answers

    Study Notes

    Gene Knockout and Genome Editing

    • Knockout mouse: a mouse in which a specific gene is knocked out in all tissues
    • ES cells: Embryonic Stem cells used to create knockout mice
    • Knockout construct can insert at the site of interest or somewhere else in the nuclear genome, which can be determined by ganciclovir sensitivity
    • LoxP-Cre recombination can be used to knockout specific genes in specific cells or tissues

    CRISPR-Cas System

    • clusters of regularly-interspaced short palindromic repeats (CRISPR) were first observed in bacteria to recognize and digest foreign DNA
    • CAS: CRISPR-associated proteins, such as nuclease Cas9, used to digest DNA
    • CRISPR RNA (crRNA) and trans-activating CRISPR RNA (tracrRNA) are used to recognize specific DNA regions
    • PAM (protospacer adjacent motifs) are important for guiding crRNA and tracrRNA to the gene of interest
    • NHEJ (Non-homologous end-joining) and HDR (Homology Directed Repair) are two repair pathways used to repair double-stranded breaks
    • Applications of CRISPR-Cas9 include genome editing, gene regulation, and future directions in human therapeutics and ecological engineering

    Gene Inactivation

    • RNA interference (RNAi) using siRNA (small interfering RNA) can be used to inactivate genes by degrading specific RNA
    • Sequencing of DNA can be done using conventional sequencing, whole-genome sequencing, whole-exome sequencing, and transcriptome sequencing

    Genetic Analysis

    • Targeted mutagenesis by homologous recombination can be used to knockout specific genes in yeast and mouse
    • Genetic complementation analysis can be used to determine whether recessive mutations are in the same or different genes
    • Analysis of double mutants can reveal the order of steps in biosynthetic pathways and interacting or redundant proteins

    DNA Cloning

    • DNA cloning involves the replication of recombinant DNA within host cells
    • Isolation, sequencing, and manipulation of purified DNA fragments can be done using DNA cloning
    • A plasmid (vector) contains sequences to maintain it in the host, such as replication origin and selection marker gene
    • Restriction enzymes, such as EcoRI, can be used to cleave palindromic sequences and create recombinant DNA

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    Related Documents

    Genetically Modified Cells and Organisms PDF

    Description

    This quiz covers genetic engineering techniques, including gene knockout, CRISPR-Cas system, and recombinase-mediated gene modification in specific cells and tissues.

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