Name: crispr gene editing basics
Uploaded: 2021-04-07T12:12:00.000Z
Duration: 1 min 59 s
Channel: JoVE

K12 Standards

north-carolina-k-12-science-standard-course-of-study

NC K-12 Science Standard Course of Study

9-12

Biology

heredity-inheritance-and-variation-of-traits

Heredity- Inheritance and Variation of Traits

LS.Bio.8.2

CRISPR

The CRISPR-Cas9 system is a DNA editing tool that stands for, Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR associated protein 9.

First observed in bacteria, CRISPR-Cas9 is a means of defense against viruses. As foreign viral DNA enters a bacterium, it&#39;s processed into smaller fragments, which may be inserted into a region of the bacterial genome called a CRISPR Locus.

When the region is transcribed, the product associates with smaller RNAs called tracrRNAs which may help to orient both the Cas9 protein and RNAse to the molecule. The latter of which cleaves the transcript.

The end result is several complexes each consisting of a Cas9 protein, tracrRNA and a crRNA derived from DNA in the Locus. The CRISPR RNA in these structures recognizes and guides Cas9 to viral DNA which is then cleaved and destroyed.

Scientists harness CRISPR-Cas9 by synthesizing individual RNA molecules that mimic tracrRNA and CRISPR RNA which can target a gene of interest. For example when two such guide RNAs are introduced into cells with Cas9 and both target the same gene, a sequence can be excised.

Once this target region is removed the cut ends are reconnected and the effects on the cells are observed.

Thus the CRISPR-Cas9 system is modified from a bacterial mechanism. And can be employed for an array of gene editing techniques.

crispr

Genome editing technologies allow scientists to modify an organism&rsquo;s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated protein 9. A basic CRISPR-Cas9 system consists of a Cas9 endonuclease and a small RNA that guides Cas9 to the target DNA.

<h4>Origin</h4>

CRISPR sequences were first observed in bacteria and later identified in archaea. Researchers discovered that the CRISPR-Cas9 system serves an adaptive immune defense against invading viruses. Many bacteria and most archaea capture short sequences of the viral DNA to create a library of virus DNA segments, or CRISPR arrays. When the prokaryotes are re-exposed to the same virus or class of viruses, CRISPR arrays are used to transcribe small RNA segments that help recognize viral invaders and subsequently destroy viral DNA with Cas9 or a similar endonuclease.

<h4>Using CRISPR-Cas9 Technology</h4>

CRISPR-Cas9 is commonly used in the laboratory to remove DNA and insert a new DNA sequence in its place. To achieve this, researchers must first create a small fragment of RNA called the guide RNA, with a short sequence called the guide sequence that binds to a specific target sequence on genomic DNA. The guide RNA can also associate with Cas9 (or other endonucleases like Cpf1). The guide RNA and Cas9 protein are administered to a cell of interest where the guide RNA identifies the target DNA sequence and Cas9 cleaves it.

The cell&rsquo;s machinery then repairs the broken strands by inserting or deleting random nucleotides, rendering the target gene inactive. Alternatively, a customized DNA sequence may be introduced into the cell along with the guide RNA and Cas9, that serves as a template for the repair machinery and replaces the excised sequence. This is a highly effective way for researchers to &ldquo;knock out&rdquo; a gene to study its effects or replace a mutated gene with a normal copy in hopes of curing a disease.

<h4>Ethical and Feasibility Considerations in Humans</h4>

As a result of the significant gene modification capabilities of the CRISPR-Cas9 system, there has been great debate over its use, especially in regards to embryo editing. A Chinese scientist recently claimed to have created genome-edited babies using CRISPR technology to disable a gene involved in HIV infection. This led to a global outcry from scientists concerned about the ethical and safety considerations of the procedure. Many have called the move premature, and others have expressed concerns over off-target genomic effects. While the number of possible biotech applications for the CRISPR-Cas9 system is numerous, it is important to consider future challenges that may arise as a result of its use.

Genome editing technologies allow scientists to modify an organism&rsquo;s DNA via the addition, removal, or rearrangement of genetic material at specific ...

Education

JoVE Core

Molecular Biology

Unit 2

Analyzing Gene Expression and Function

KEY TERMS AND CONCEPTS

crispr gene editing basics

CRISPR is a gene editing tool that helps scientists make targeted changes to DNA. It uses a guide system to find a chosen sequence and a cutting protein to act at that spot.

This method is based on a natural defense system found in bacteria. In high school biology, it is often studied as a way to see how genes can be edited with precision.

CRISPR is widely used in molecular biology research. It helps scientists explore how genes function and how DNA changes can affect living things.

CRISPR Gene Editing Basics

in vitro mutagenesis

In Vitro Mutagenesis for Gene Analysis

In vitro mutagenesis is a lab method used to change DNA and study gene function. It helps scientists test how specific DNA changes affect a gene’s activity and the traits it controls. A common approach is to make a known change in a DNA sequence and then examine the result. This can be done by deleting, inserting, or altering selected bases in a gene. The method is useful for linking a DNA change to a biological effect. Researchers use in vitro mutagenesis in molecular biology, genetics, and...

genetic screens

Using Genetic Screens to Find Disease Genes

Genetic screens are tools used to find genes and mutations linked to specific traits. They are also used to identify people or groups at risk for genetic diseases. That can support early intervention, targeted therapy, and reproductive choices. Forward genetic screens, also called classical screens, start with random DNA changes in an organism. Researchers may use radiation, mutagens, or added bases to create mutations that lead to visible phenotype changes. The mutant is then inbred so...

test cross

Test Cross in Genetic Inheritance

A test cross is a genetics tool used to determine the genotype of an organism with a dominant trait. It helps reveal whether the dominant individual is homozygous or heterozygous by crossing it with a recessive individual. The outcome of the cross shows the hidden genotype. If any offspring show the recessive trait, the dominant parent must carry one recessive allele. If all offspring show the dominant trait, the dominant parent is likely homozygous dominant. Test crosses are common in basic...

complementation tests

Using Crosses to Group Mutations

A complementation test uses a simple cross to find out whether two mutations are in the same gene or in different genes. It helps show how mutations are arranged on chromosomes. Edward Lewis first performed this test in the 1940s while studying fruit flies. To do the test, organisms heterozygous for different mutations are crossed pairwise in all combinations. If the mutations are in different genes, they can complement each other. In that case, each mutation provides the missing genetic...

single nucleotide polymorphisms-snps

SNPs as Genetic Markers and Variants

Single nucleotide polymorphisms, or SNPs, are small DNA changes at one specific genomic position. They are the most common type of sequence variation in the human genome. A point mutation counts as an SNP when it appears in more than 1% of the population.

bacterial transformation

Bacterial Transformation in the Lab

Bacterial transformation is a lab method used to move new DNA into bacteria. Scientists use it to change how bacteria behave or to help them carry a gene of interest. In high school biology, this process is often shown with plasmids, which are small circular DNA molecules. During transformation, bacteria take up DNA from their surroundings. The new DNA can then be used by the cell, depending on the experiment. This technique is an important part of genetic engineering and is widely used in...

transgenic organisms

Making Transgenic Organisms

Transgenic organisms are living things with DNA added from another organism. A transgene is the inserted gene. Scientists use this method to change an organism’s traits and to study how genes work. To make a transgenic organism, researchers first identify the gene they want to add. They then place that gene into the target organism’s DNA. Once the gene is integrated, the organism can pass it to new cells as it grows. This approach is useful in biology because it helps scientists analyze gene...

reproductive cloning

Cloning an Organism by Reproduction

Reproductive cloning creates a new organism that is genetically the same as the donor. It is different from cloning genes or cells because the goal is to make a whole living individual. This process usually relies on somatic cell nuclear transfer. In this method, the nucleus from a body cell is placed into an egg cell that has had its own nucleus removed. The egg can then develop into an embryo with DNA from the donor cell. Reproductive cloning is used to produce a copy of an organism, such...

CRISPR is a gene editing tool that helps scientists make targeted changes to DNA. It uses a guide system to find a chosen sequence and a cutting protein to act at that spot. This method is based on a natural defense system found in bacteria. In high school biology, it is often studied as a way to see how genes can be edited with precision. CRISPR is widely used in molecular biology research. It helps scientists explore how genes function and how DNA changes can affect living things.

experimental rnai

RNAi for Gene Silencing and Discovery

RNA interference (RNAi) is a gene-silencing process used to reduce gene expression. It can block transcription or trigger RNA degradation. Scientists first discovered this mechanism in plants in 1998 through Andrew Fire and Craig Mello. RNAi has since been found in many eukaryotes, including protozoa, flies, nematodes, insects, parasites, and mammals. Researchers use it as a practical tool to study gene function. It can show what a gene does without permanently changing the organism’s DNA. <br...

reporter genes

Reporter Genes in Cells and Plasmids

Reporter genes are protein-coding genes used to track gene activity in cells. They are often tagged to a gene of interest so scientists can see whether that gene is present or active. When the tagged genes are expressed together, reporter genes can produce visible signals such as fluorescence or luminescence. These signals help show the presence or absence of a gene of interest in an organism. They can also reveal the pattern of gene expression or the location of a DNA segment or protein...

in-situ hybridization

ISH Probes for DNA and RNA Detection

In situ hybridization (ISH) uses labeled probes to detect and localize specific DNA or RNA molecules in cells, tissues, or tissue sections. The probe is a complementary strand of DNA or RNA that binds to a matching nucleotide sequence in the sample. Since its first use in 1969 for nucleic acid studies, ISH has become an important tool in research and in clinical diagnosis. Several probe types can be used in ISH. These include single-stranded DNA probes, double-stranded DNA probes, antisense...

chromatin immunoprecipitation- chip

ChIP Methods for DNA-Protein Binding

Chromatin immunoprecipitation, or ChIP, is an antibody-based method for finding where proteins bind to DNA. It is used to identify DNA sites bound by transcription factors or histone proteins. ChIP also helps show which histone changes are present, including acetylation, phosphorylation, and methylation. ChIP is commonly divided into two forms: X-ChIP and N-ChIP. X-ChIP first cross-links proteins to DNA inside the cell, then breaks the DNA into pieces by sonication. The protein-DNA complexes...

synthetic biology

Synthetic Biology in Crops and Cells

Synthetic biology combines engineering, molecular biology, cell biology, and systems biology to redesign living systems. Scientists use it to remodel organisms from nature or to build new synthetic organisms. These tools can support protein or enzyme production, bioremediation, the production of useful macromolecules, and the addition of helpful traits to crops. One well-known example is golden rice. This genetically modified rice plant makes grains rich in β-carotene, which is a precursor of...

ribosome profiling

Ribosome Profiling for Translation Snapshot

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transgenic plants

Transgenic Plants for Better Crops

Transgenic plants are crops that have been changed with recombinant DNA technology, also called transgenesis. This method adds a foreign gene or removes a harmful one from an organism. The result is a genetically modified organism. The first transgenic plant was a tobacco plant made in 1983, and it was resistant to tobacco mosaic virus. Since then, transgenic plants have been developed for many uses in agriculture, horticulture, and ornamental plants. Some are designed to resist pests or...

gene therapy

Gene Therapy for Disease Treatment

Gene therapy is a treatment approach that changes gene activity to help address disease. It focuses on the genetic cause of a problem rather than only easing symptoms. In this way, it is used in disease intervention. One common strategy is to replace a faulty gene with a normal copy. Another strategy is to inactivate a gene that is causing harm. Gene therapy can also introduce a new gene to help cells work more normally.

epistasis analysis

Gene Interaction Maps in Epistasis

Gene interaction maps in epistasis show how different genes work together to shape a phenotype. In epistasis, the interaction of genes or alleles at different locations affects the final trait. This often happens when one gene masks or interferes with the expression of another gene. Epistasis is common when genes are part of the same biochemical pathway. In that case, the expression of one gene can depend on a gene product made earlier in the pathway. Genes that mask or inhibit other genes in...

genome-wide association studies-gwas

Using SNP Patterns to Link Disease Risk

Genome-wide association studies, or GWAS, look for common SNPs, or single nucleotide polymorphisms, that may be linked to disease. Researchers compare how often a SNP appears in people with a disease and in people without it. When a SNP is seen more often in the disease group, it is considered associated with that disease. GWAS uses chi-square analysis to test the chance that an allele is linked to the disease. This approach does not require scientists to first find the target gene behind the...

key terms and concepts

Explore CRISPR as a gene editing tool, its bacterial origins, and how it is used in molecular biology to study DNA and gene function.

CRISPR Gene Editing Basics | JoVE K12

molecular biology

JoVE Core Molecular Biology showcases the molecular processes of the cell through clear and concise animated video lessons. Additionally, in scientist-in-action videos, these concepts' application is demonstrated through real experiments in modern laboratories.

Tennessee Academic Standards

High School

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Links Among Engineering, Technology, Science, and Society

BIO1.ETS2.1

Virginia  Standards of Learning

HIgh School

BIO.5

BIO.5.e

Arizona Science Standards

Life Science

HS.L3

HS.L3U3.26

Alabama Course of Study Science

from-molecules-to-organisms-structures-and-processes

From Molecules to Organisms: Structures and Processes

9-12.B.1.2.c

Florida Department of Education

SC.912.L.16

SC.912.L.16.7

myJoVE Corporation

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LS.Bio.6.2

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Dihybrid Crosses

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Law of Independent Assortment

Chi-square Analysis

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Multiple Allele Traits

Incomplete Dominance

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Lethal Alleles

Agouti: A Lethal Allele
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Polygenic Traits

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Background and Environment Affect Phenotype

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LS.Bio.7.1

Overview

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Probability Laws

Overview

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The Concept of Multiple Allelism

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Analyze and interpret data to explain how polygenic traits result in a wide range of phenotypes.

LS.Bio.7.2

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LS.Bio.7.3

PCR: The Polymerase Chain Reaction

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DNA Isolation

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Restriction Enzymes

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Analyze and interpret data to compare DNA samples.

LS.Bio.8.1

Understand genetic mechanisms for variation.

CRISPR Gene Editing Basics

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