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

Das CRISPR-Cas9-System ist ein DNA-Bearbeitungswerkzeug, das für geclusterte, regelmäßig verteilte kurze palindromische Wiederholungen und CRISPR-assoziiertes Protein 9 steht.
 CRISPR-Cas9 wurde erstmals in Bakterien beobachtet und ist ein Mittel zur Abwehr von Viren.
 Wenn fremde virale DNA in ein Bakterium eindringt, wird sie zu kleineren Fragmenten verarbeitet, die in eine Region des Bakteriengenoms, den CRISPR-Locus, eingefügt werden können.
 Wenn die Region transkribiert wird, assoziiert sich das Produkt mit kleineren RNAs, die tracrRNAs genannt werden.
 Dies kann dazu beitragen, sowohl das Cas9-Protein als auch die RNAse an dem Molekül zu orientieren, das das Transkript spaltet.
 Das Endergebnis sind mehrere Komplexe, die jeweils aus einem Cas9-Protein, TracrRNA und einer CRISPR-RNA bestehen, die von DNA im Locus abgeleitet sind.
 Die CRISPR-RNA in diesen Strukturen erkennt und führt Cas9 zu viraler DNA, die dann gespalten und zerstört wird.
 Wissenschaftler nutzen CRISPR-Cas9, indem sie einzelne RNA-Moleküle synthetisieren, die TracrRNA und CrRNA imitieren, die auf ein Gen von Interesse abzielen können.
 Wenn zum Beispiel zwei solche Leit-RNAs mit Cas9 in Zellen eingeführt werden und beide auf dasselbe Gen abzielen, kann eine Sequenz herausgeschnitten werden.
 Sobald dieser Zielbereich entfernt ist, werden die Schnittenden wieder verbunden und die Auswirkungen auf die Zellen beobachtet.
 Somit ist das CRISPR-Cas9-System durch einen Bakterienmechanismus modifiziert und kann für eine Reihe von Gen-Editing-Techniken eingesetzt werden.

Thurtle-Schmidt, Deborah M. und Te-Wen Lo. "Molekularbiologie am Puls der Zeit: Ein Überblick über CRISPR/CAS9-Gen-Editing für Studenten." Biochemie und molekularbiologische Bildung 46, Nr. 2 (2018): 195–205. [<a href="https://doi.org/10.1002/bmb.21108" target="__blank">Quelle</a>]

Lander, Eric S. "Die Helden von CRISPR." Zelle 164, Nr. 1 (14. Januar 2016): 18–28. [<a href="https://doi.org/10.1016/j.cell.2015.12.041" target="__blank">Quelle</a>]

crispr

Genom-Editierung Techniken erlauben es Wissenschaftlern, die DNA eines Organismus zu verändern durch das Hinzufügen, das Entfernen oder der Neuordnung von genetischem Material an spezifischen Genom loci. Diese Techniken könnten wahrscheinlich zur Heilung von genetischen Krankheiten wie Hämophilie und Sichelzellanämie eingesetzt werden. Ein beliebtes und weit verbreitetes Forschungsinstrument zur DNA-Editierung, dass zu einer sicheren und effektiven Behandlungsmethode für genetische Krankheiten führen könnte, ist das CRISPR-Cas9-System. CRISPR-Cas9 steht für Clustered Regularly Interspaced Short Palindromic Repeats und CRISPR-associated Protein 9. Das grundlegende CRISPR-Cas9-System besteht aus einer Cas9-Endonuclease und einer kleinen RNA, die Cas9 zu dir Ziel-DNA führt.
 
 <h4>Die Herkunft</h4>
 
 CRISPR-Sequenzen wurden zuerst in Bakterien beobachtet und später in Archaeen identifiziert. Die Forscher entdeckten, dass das CRISPR-Cas9-System einer adaptiven Immunabwehr gegen eindringende Viren dient. Viele Bakterien und die meisten Archaeen fangen kurze Sequenzen der Virus-DNA ein, um eine Bibliothek von Virus-DNA-Segmenten, sogenannte CRISPR-Arrays, zu erstellen. Wenn Prokaryoten erneut dem gleichen Virus oder der gleichen Virusklasse ausgesetzt werden, werden diese CRISPR-Arrays verwendet, um kleine RNA-Segmente zu transkribieren. Diese helfen dabei, virale Eindringlinge zu erkennen und anschließend die virale DNA mit Cas9 oder einer ähnlichen Endonuklease zu zerstören.
 
 <h4>Verwendung der CRISPR-Cas9 Technologie</h4>
 
 Das CRISPR-Cas9 wird regulär im Labor verwendet, um DNA zu entfernen und eine neue DNA-Sequenz an dieser Stelle einzufügen. Um dies zu erreichen, müssen die Forscher zunächst ein kleines RNA-Fragment (spacer-RNA) mit einer kurzen Sequenz (spacer-Sequenz) erstellen, die an eine bestimmte Zielsequenz in der genomischen DNA bindet. Die Spacer-RNA kann auch Cas9 (oder anderen Endonukleasen wie Cpf1) binden. Die Spacer-RNA und das Cas9-Protein werden in die gewünschte Zelle eingeführt, wo dann die spacer-RNA die Ziel-DNA-Sequenz identifiziert und Cas9 sie letztlich spaltet.
 
 Die Maschinerie der Zelle repariert dann die gebrochenen DNA-Stränge durch Einfügen oder Entfernen von zufälligen Nucleotiden. Dadurch wird das Zielgen inaktiviert. Alternativ kann eine spezifische DNA-Sequenz zusammen mit der spacer-RNA und Cas9 in die Zelle eingeführt werden. Diese dient als Vorlage für die Reparaturmaschinerie und ersetzt die entfernte Sequenz. Diese Methode ist hochwirksam, um bestimmte Gene “auszuschalten”, zu untersuchen oder ein mutiertes Gen durch eine normale Kopie zu ersetzen, in der Hoffnung, eine Krankheit heilen zu können.
 
 <h4>Ethische und praktische Erwägungen beim Menschen</h4>
 
 Aufgrund der signifikanten gentechnischen Möglichkeiten des CRISPR-Cas9-Systems gab es eine große Debatte hinsichtlich seiner Verwendung. Besonders wurde die Verwendung des CRISPR-Cas9-Systems im Hinblick auf die Modifizierung von Embryos diskutiert. Ein chinesischer Wissenschaftler behauptete kürzlich, genomeditierte Babys mit Hilfe der CRISPR-Technologie erzeugt zu haben. Dabei behauptete er ein bei HIV-Infektionen beteiligtes Gen deaktiviert zu haben. Dies führte zu einem weltweiten Aufschrei von Wissenschaftlern, die über die ethischen und sicherheitstechnischen Aspekte des Verfahrens besorgt sind. Viele haben den Schritt als verfrüht bezeichnet, während andere äußerten ihre Bedenken zu möglichen genomischen Off-Target-Effekten. Obwohl die Zahl der möglichen biotechnologischen Anwendungen für das CRISPR-Cas9-System zahlreich ist, sollte man zukünftige Herausforderungen berücksichtigen, die sich aus dem Einsatz des Systems ergeben können.

Genom-Editing-Technologien ermöglichen es Wissenschaftlern, die DNA eines Organismus durch Hinzufügen, Entfernen oder Umordnen von genetischem Material zu bestimmten Zeitpunkten zu verändern.

Education

Lehre

JoVE Core

Molecular Biology

Molekularbiologie

Unit 2

Einheit 2

Analyzing Gene Expression and Function

Analyse von Genexpression und -funktion

KEY TERMS AND CONCEPTS

KERNBEGRIFFE UND KONZEPTE

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

Ribosome profiling is a deep sequencing method that gives a snapshot of active translation in a cell. It sequences the mRNAs protected by ribosomes. This shows which messages are being read to make proteins at a specific moment. The method has several uses in biology. It can track translation in a living organ or tissue type. It also helps measure new protein synthesis levels. Researchers use it to find translated open reading frames, which are stretches of RNA that can make protein. It can...

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 veranschaulicht die molekularen Prozesse der Zelle in prägnanten animierten Video-Lektionen. In Videos von Wissenschaftlern in Aktion wird die Anwendung dieser Konzepte zusätzlich durch reale Experimente in modernen Laboren demonstriert.

Tennessee Academic Standards

High School

links-among-engineering-technology-science-and-society

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

What is Meiosis?

Meiosis is the process by which diploid cells divide to produce haploid daughter cells. In humans, each diploid cell contains 46 chromosomes, half from ...

Meiosis I

Meiosis II

Meiosis vs. Mitosis

Crossing Over

Crossing over is the exchange of genetic information between homologous chromosomes during prophase I of meiosis I. Genetic recombination gives rise to ...

Nondisjunction

Nondisjunction is the failure of homologous chromosomes or sister chromatids to separate correctly and move to the opposite poles of the cells. This ...

Spermatogenesis

Spermatogenesis is the process by which haploid sperm cells are produced in the male testes. It starts with stem cells located close to the outer rim of ...

Oogenesis

In human women, oogenesis produces one mature egg cell or ovum for every precursor cell that enters meiosis. This process differs in two unique ways from ...

Fertilization

During fertilization, an egg and sperm cell fuse to create a new diploid structure. In humans, the process occurs once the egg has been released from the ...

Cleavage and Blastulation

After a large-single-celled zygote is produced via fertilization, the process of cleavage occurs while zygotes travel through the uterine tube. Cleavage ...

Gastrulation

Gastrulation establishes the three primary tissues of an embryo: the ectoderm, mesoderm, and endoderm. This developmental process relies on a series of ...

Neurulation

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Cell Migration

Cell migration, the process by which cells move from one location to another, is essential for the proper development and viability of organisms ...

Determination

During embryogenesis, cells become progressively committed to different fates through a two-step process: specification followed by determination. ...

Use models to explain how DNA is passed from parents to offspring through the processes of meiosis and fertilization in sexual reproduction.

LS.Bio.6.1

Mutations

Mutations are changes in the sequence of DNA. These changes can occur spontaneously or they can be induced by exposure to environmental factors. Mutations ...

genome-size-and-the-evolution-of-new-genes

Genome Size and the Evolution of New Genes

types-of-genetic-transfer-between-organisms

Types of Genetic Transfer Between Organisms

Base-pairing and DNA Repair

The DNA Replication Fork

Lagging Strand Synthesis

The Replisome

Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template ...

Replication in Prokaryotes

DNA replication has three main steps: initiation, elongation, and termination. Replication in prokaryotes begins when initiator proteins bind to the ...

Replication in Eukaryotes

In eukaryotic cells, DNA replication is highly conserved and tightly regulated. Multiple linear chromosomes must be duplicated with high fidelity before ...

Overview of DNA Repair

Base Excision Repair

Nucleotide Excision Repair

DNA Distortion and Damage
Cells are regularly exposed to mutagens&#8212;factors in the environment that can damage DNA and generate mutations. UV ...

Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the ...

Fixing Double-strand Breaks

Homologous Recombination

Gene Conversion

Restarting Stalled Replication Forks

Construct an explanation to summarize how inheritable genetic variations may result from: new genetic combinations in meiosis, mutations during replication, or mutations caused by environmental factors.

LS.Bio.6.2

An Overview of Genetic Analysis

An organism&#8217;s physical traits, or phenotype, are a product of its genotype, which is the combination of alleles (gene variants) inherited from its ...

Genetic Crosses

To dissect genetic processes or create organisms with novel suites of traits, scientists can perform genetic crosses, or the purposeful mating of two ...

Punnett Squares

Monohybrid Crosses

Dihybrid Crosses

Trihybrid Crosses

Trihybrid Crosses
Some of Mendel&#8217;s crosses examined three pairs of contrasting characteristics. Such a cross is called a trihybrid cross. A ...

Law of Independent Assortment

Chi-square Analysis

The chi-square test is a statistical hypothesis test. It is used to check whether there is a significant difference between an expected value and an ...

Pedigree Analysis

Multiple Allele Traits

Incomplete Dominance

Gregor Mendel&#39;s work (1822 - 1884) was primarily focused on pea plants. Through his initial experiments, he determined that every gene in a diploid ...

Lethal Alleles

Agouti: A Lethal Allele
Lucien Cu&#233;not discovered lethal alleles in 1905 while studying the inheritance of coat color in mice. The agouti gene is ...

Polygenic Traits

background-and-environment-affect-phenotype

Background and Environment Affect Phenotype

Although the genetic makeup of an organism plays a major role in determining the phenotype, there are also several environmental factors, such as ...

Use mathematics and computational thinking to predict the variation and distribution of expressed traits based on: Mendelian inheritance, co-dominance, incomplete dominance, multiple alleles, and sex-linked inheritance.

LS.Bio.7.1

Overview

A pedigree is a diagram displaying a family&#x2019;s history of a trait. Analyzing pedigrees can reveal (1) whether a trait is dominant or ...

Probability Laws

Overview

The probability of inheriting a trait can be calculated using the sum and product rules. The sum rule is used to calculate the probability of ...

The Concept of Multiple Allelism

Multiple allelism describes genes that exist in three or more allelic forms. Although diploid organisms, like humans, ...

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

LS.Bio.7.2

Pleiotropy

Pleiotropy is the phenomenon in which a single gene impacts multiple, seemingly unrelated phenotypic traits. For example, defects in the SOX10 gene cause ...

Nature and Nurture

Many human characteristics, like height, are shaped by both nature&mdash;in other words, by our genes&mdash;and by nurture, or our environment. For ...

Construct an explanation to summarize how traits result from interactions of genetic factors (multiple genes and/or alleles) and environmental factors.

LS.Bio.7.3

PCR: The Polymerase Chain Reaction

The polymerase chain reaction, or PCR, is a technique used to amplify DNA through thermocycling &#8211; cyles of temperature changes at fixed time ...

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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