Name: crispr-cas9 gene editing in action
Uploaded: 2019-03-11T09:21:22.000Z
Duration: 1 min 59 s
Channel: JoVE

K12 Standards

Tennessee Academic Standards

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Biology

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Links Among Engineering, Technology, Science, and Society

BIO1.ETS2.3

CRISPR

CRISPR-Cas9系统 是一种DNA编辑工具， 代表有规则的间隔排列的短回文重复序列 和CRISPR相关蛋白9。 首先在细菌中观察到， CRISPR-Cas9是一种防御病毒的手段。 当外来病毒DNA进入细菌时， 会被加工成更小的片段。
该基因可能被插入细菌基因组中 一个被称为CRISPR基因座的区域。 当该区域被转录时， 该产物与被称为转录核糖核酸的较小核糖核酸结合， 这可能有助于将Cas9蛋白和 核糖核酸酶定位于该分子。 后者切割了转录本。
最终的结果是几个复合物， 每个复合物由一个Cas9蛋白、四氢核糖核酸和一个 来源于基因座的脱氧核糖核酸组成。 这些结构中的CRISPR核糖核酸识别 并引导Cas9进入 病毒的脱氧核糖核酸，然后被切割和破坏。 科学家通过合成单个的 核糖核酸分子 来控制CRISPR-Cas9，这些核糖核酸分子模仿 能靶向感兴趣基因的tracrRNA和CRISPR核糖核酸。
例如，当两个这样的引导核糖核酸 被引入到具有Cas9的细胞中， 并且两者都靶向相同的基因时， 序列可以被切除。 一旦该目标区域被移除， 切割端被重新连接， 并且观察对细胞的影响。 因此CRISPR-Cas9系统是由 细菌机制改造而来的。 并且可以用于一系列基因编辑技术。

Thurtle-Schmidt、Deborah M. 和 Te-温 Lo. "前沿分子生物学:本科生 CRISPR/CAS9 基因编辑综述"。生物化学与分子生物学教育 46，第 2 期（2018）:195-205。[<a href="https://doi.org/10.1002/bmb.21108" target="__blank">来源</a>]

兰德，埃里克 S. "CRISPR 的英雄"。单元格 164，第 1 期（2016 年 1 月 14 日）:18-28。[<a href="https://doi.org/10.1016/j.cell.2015.12.041" target="__blank">来源</a>]

基因编辑技术

基因组编辑技术允许科学家通过在特定基因组位置添加、移除或重新排列遗传物质来修改生物体的DNA。这些技术有可能被用于治疗血友病和镰状细胞性贫血等遗传疾病。crispr-cas9系统是一种广受欢迎、应用广泛的DNA编辑研究工具，可以安全有效地治疗遗传性疾病。crispr-cas9代表簇状规则间隔的短回文重复序列和crispr相关蛋白9。一个基本的CRISPR-CAS9系统由一个CAS9内切酶和一个引导CAS9到达目标DNA的小RNA组成。

<h4>起源</h4>
 
CRISPR 序列最初在细菌中观察到，后来在古细菌中鉴定出来。研究人员发现CRISPR-Cas9 系统对入侵病毒具有适应性免疫防御作用。许多细菌和大多数古细菌捕获病毒DNA的短序列，以创建病毒DNA片段库或CRISPR 阵列。当原核生物再次暴露于同一病毒或同一类病毒时， CRISPR阵列被用来转录小RNA片段，帮助识别病毒侵略者，然后用Cas9或类似的核酸内切酶破坏病毒DNA。
 
<h4>使用CRISPR-Cas9技术</h4>
 
CRISPR-Cas9是实验室常用的去除DNA并插入新的DNA序列的方法。为了实现这一目标，研究人员必须首先创建一个称为导向RNA的RNA小片段，其短序列称为导向序列，与基因组DNA上的特定靶序列结合。引导RNA还可以与 Cas9 (或Cpf1 等其它核酸内切酶)结合。引导RNA和Cas9 蛋白被注射到兴趣的细胞中，引导RNA识别目标DNA序列， Cas9将其切割。
 
然后，细胞的机器通过插入或删除随机核苷酸来修复断裂的链，从而使目标基因失去活性。或者，可以将定制的DNA序列与作为修复机器的模板的引导RNA和Cas9一起引入细胞，并替换切除的序列。对于研究人员来说，这是一种非常有效的方法，可以“敲除”一个基因来研究其作用，或者用一个正常的拷贝替换一个突变的基因，以期治愈一种疾病。
 
<h4>人类的伦理和可行性考虑</h4>
 
由于CRISPR-Cas9 系统具有显著的基因修饰能力，人们对它的使用，特别是在胚胎编辑方面，进行了激烈的争论。一位中国科学家最近声称，他们利用 CRISPR技术创造了基因编辑婴儿，使一种与艾滋病病毒感染有关的基因失效。这导致了全球科学家对该程序的伦理和安全考虑的强烈抗议。许多人认为此举为时过早，其他人则对非靶基因效应表示担忧。尽管CRISPR-Cas9系统可能的生物技术应用数量众多，但重要的是要考虑其使用可能带来的未来挑战。

基因组编辑技术使科学家能够通过在特定位置添加、移除或重排遗传物质，对生物体的DNA进行修饰。

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生物学

Genetics

遗传学

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KEY TERMS AND CONCEPTS

关键术语与概念

crispr-cas9 gene editing in action

CRISPR-Cas9 is a genome editing tool that lets scientists change DNA at a specific location. It can add, remove, or rearrange genetic material. Researchers study it because it may help treat genetic disorders such as hemophilia and sickle cell anemia.

CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats, and Cas9 is a CRISPR-associated protein 9. The basic system includes a Cas9 endonuclease, which is an enzyme that cuts DNA, and a small RNA that guides Cas9 to the target sequence. Together, they make CRISPR-Cas9 a precise research tool for DNA editing.

CRISPR sequences were first seen in bacteria and later found in archaea. Scientists discovered that this system acts like an adaptive immune defense against viruses. Many bacteria and most archaea store short pieces of viral DNA in CRISPR arrays. When the same virus returns, these RNA segments help identify it, and Cas9 or a similar endonuclease destroys the viral DNA.

In the lab, CRISPR-Cas9 is often used to remove DNA and replace it with a new sequence. Researchers first design a guide RNA, which contains a guide sequence that matches a target sequence on genomic DNA. The guide RNA and Cas9 are then placed in the cell of interest. The guide RNA finds the target, Cas9 cuts the DNA, and the cell repairs the break by adding or deleting random nucleotides, which can turn the gene off.

Scientists can also add a custom DNA template with the guide RNA and Cas9. The repair machinery can use that template to replace the removed sequence. This makes CRISPR useful for gene knockouts and for replacing a mutated gene with a normal copy in hopes of treating disease. Other endonucleases, such as Cpf1, can also work with guide RNA.

CRISPR-Cas9 has also raised major ethical and safety questions, especially about embryo editing. A Chinese scientist claimed to have used CRISPR to create genome-edited babies by disabling a gene linked to HIV infection. That claim led to global concern because many scientists thought the work was premature and worried about off-target genomic effects. CRISPR has many possible biotech uses, but future challenges still need careful attention.

CRISPR-Cas9 Gene Editing in Action

what is genetic engineering?

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crispr

CRISPR-Cas9 is a genome editing tool that lets scientists change DNA at a specific location. It can add, remove, or rearrange genetic material. Researchers study it because it may help treat genetic disorders such as hemophilia and sickle cell anemia. CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats, and Cas9 is a CRISPR-associated protein 9. The basic system includes a Cas9 endonuclease, which is an enzyme that cuts DNA, and a small RNA that guides Cas9 to the...

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genomics

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Genomics studies the full set of genetic material in an organism. In humans, that genome includes the DNA carried in 23 pairs of chromosomes in the nucleus, along with mitochondrial DNA. Genomics looks at both coding DNA and non-coding DNA, which helps scientists study how living things are built and how they differ. This field is useful in many areas of biology and medicine. Scientists use genomics to build phylogenetic trees, improve crop productivity and sustainability, help with criminal...

key terms and concepts

Learn how CRISPR-Cas9 edits DNA, uses guide RNA and Cas9, and why scientists debate its use in embryos and gene therapy.

CRISPR-Cas9 Gene Editing in Action | JoVE K12

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JoVE Core系列通过300多节动画视频课简明易懂地讲解生物学中的关键概念，以及150余段视频展示科学家在现代实验室开展实际研究实验，让生物学变得平易近人。

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EXPLANATION: The utilization of new technologies in any field of science is dependent on both economic and social factors. In addition to evaluating these factors, scientists must also consider long-term consequences that may not be initially apparent. The emphasis should be on the construction of a rationale argument that supports a position on the use of an application with ethical and social impact. Students should begin to appreciate the differences in ethical values that exist, and recognize that discussion of these values is imperative as knowledge and technology continue to advance, even when resolutions of differences can be rare. After investigation of a specific biotechnology application, students can write a position paper and/or participate in a classroom debate.

CRISPR-Cas9 Gene Editing in Action

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