Name: recombinant dna and gene cloning
Uploaded: 2021-04-07T12:11:59.000Z
Duration: 1 min 9 s
Channel: JoVE

K12 Standards

Tennessee Academic Standards

High School

Biology

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

Links Among Engineering, Technology, Science, and Society

BIO1.ETS2.1

Recombinant DNA

재조합된 DNA는여러 출처, 보통 다른 종에서 온 DNA로실험실에서 조합이 되어임상 및 과학 연구에서 널리 사용되고 있다재조합된 DNA는 보통 관심 유전자로 구성되는데여기서는 기부자 유기체에서 나온 인슐린으로벡터에 투입이 되는데벡터는 바이러스, 플라스미드 같은다른 유기체에서 온 자기 복제 DNA로서 박테리아 DNA의작은 원형의 조각이다각 출처의 DNA는 제한적 핵산 분해 효소로먼저 잘리는데, 이 효소는DNA를 특정의 서열로 쪼개는 효소이다기부자 DNA는 DNA 연결 효소의 도움으로벡터 DNA의 끝에 연결이 된다재조합된 DNA는 박테리아나 효모 같은 숙주 유기체에넣어지고, 거기서 복제가 되고관심 유전자가 많이 복제된다그 유전자는 그 다음에 더 쉽게 연구가 되거나인슐린의 경우처럼숙주의 세포 시스템을 이용하여 단백질을 만들도록 할 수 있다

칸, 술리만, 무하마드 와지드 울라, 라베아 시디크, 굴람 나비, 세리시 마난, 무하마드 유사프, 홍웨이 호우. 2016. "삶의 질을 향상시키는 재조합 DNA 기술의 역할." 2016 국제유전체학회지. [<a href="https://doi.org/10.1155/2016/2405954" target="__blank">소스</a>]

마츠무라, 이치로. 2015. "Johnny가 클론을 할 수 없는 이유: 일반적인 함정과 그리 흔하지 않은 솔루션." 바이오테크닉스 59 (3): IV–XIII. [<a href="https://doi.org/10.2144/000114324" target="__blank">소스</a>]

재조합 dna

<h4>개요</h4>
 
 과학자들은 실험실에서 다른 출처, 다른 종의 DNA를 결합하여 재조합 DNA(recombinant DNA)를 만듭니다. DNA 클로닝(DNA cloning)은 과학자들이 박테리아와 같이 쉽게 조작할 수 있는 세포에 유전자를 삽입해 특정한 유전자를 연구할 수 있게 해줍니다. 재조합 DNA를 포함하는 유기체는 유전자 변형 유기체로 알려져 있습니다. 재조합 DNA 기술은 과학, 의학, 그리고 농업에 도움이 될 수 있는 새로운 유전자를 가진 유기체를 생산합니다.
 
 <h4>과학자들은 어떻게 재조합 DNA를 만들 수 있습니까?</h4>
 
 재조합 DNA의 생성은 DNA 복제와 단백질 발현을 위해 외래 DNA를 숙주 세포로 운반하는 매개체인 벡터(vector)에 관심 유전자(gene of interest)를 삽입해 이루어집니다. 가장 일반적으로 사용되는 클로닝 벡터는 숙주의 염색체 DNA와 독립적으로 복제되는 작은 원형 DNA 조각인 플라스미드입니다.
 
 재조합 DNA를 만들기 위해 관심 유전자를 포함한 공여 DNA(donor DNA)와 벡터를 제한 효소(restriction enzyme)를 사용하여 제한 부위(restriction site)라는 특정 뉴클레오타이드 서열(nucleotide sequence)에서 절단합니다. 이어서 DNA 연결효소(DNA ligase)를 사용해 관심 있는 유전자와 플라스미드가 연결되는 당-인산 뼈대를 봉합합니다.
 
 그 결과, 재조합 DNA 분자는 삽입체(insert; 인서트)라고 불리는 공여 DNA 조각이 결합한 벡터로 구성됩니다. 그런 다음 과학자들은 이 혼합 DNA 분자를 숙주 유기체(일반적으로 박테리아나 효모)에 도입하여 쉽고 빠르게 복제할 수 있습니다. 이것은 과학 연구와 다른 응용 분야에 필요한 관심 유전자의 많은 사본을 만듭니다. 또한 숙주를 이용해 이 유전자를 인간의 인슐린과 같은 원하는 단백질로 전사(transcription)하고 번역(translation)할 수 있습니다.
 
 재조합 DNA를 만드는 것은 불완전한 과정이며 종종 오류가 발생합니다. 예를 들어, 삽입체 없이 벡터가 닫히거나 삽입체가 정확하지 않을 수 있습니다(예: 거꾸로 삽입). 따라서 추가적인 연구를 위해 재조합 DNA를 사용하기 전에 오류를 확인해야 합니다. 뉴클레오타이드 서열분석(nucleotide sequencing)은 올바른 삽입체를 가진 플라스미드를 포함하는 박테리아 집락(colony)을 식별하는 데 도움이 될 수 있습니다.
 
 과학자들은 유전자와 단백질을 연구하기 위해 재조합 DNA를 사용합니다
 
 재조합 DNA 기술은 관심 있는 유전자나 단백질 생성물의 많은 사본이 필요할 때 특히 유용합니다. 그러나 어떤 연구에는 원하는 단백질의 검출이나 정제(purification) 같은 보다 복잡한 실험을 추가해야할 수도 있습니다. 이를 위해서 유전자 생성물을 식별하는 데 사용되는 단백질인 표지 단백질(protein tag)이나 리포터(reporter)를 원하는 단백질에 부착하여 융합 유전자(fusion gene) 또는 키메릭 유전자(chimeric gene)를 만들 수 있습니다.
 
 <h4>의학 및 농업 분야에서의 응용</h4>
 
 과학자들은 처음에 박테리아에서 인슐린을 생산하기 위해 재조합 DNA 기술을 사용했고, 그 결과 당뇨병을 치료할 수 있었습니다. 이 최초 발견 이후, 과학자들은 치료 목적으로 다른 재조합 DNA를 만들어냈습니다. 재조합 박테리아는 성장 호르몬 결핍 환자를 치료하기 위해 정상적인 성장과 발육에 필요한 단백질인 인간 성장 호르몬을 만듭니다. 인간과 햄스터에서 파생된 재조합 포유류 세포는 혈우병 환자를 치료하는 데 필요한 단백질인 VIII인자를 생산합니다. 재조합 DNA 기술은 필수 단백질의 대규모 생산을 위한 강력한 도구입니다.
 
 재조합 DNA 기술을 통한 농업 발전 또한 인간의 안녕에 영향을 미칩니다. 예를 들어, 옥수수 농부들은 유럽옥수수좀(European corn borer) 때문에 상당한 농작물 피해를 보았습니다. 이에 대응하여 과학자들은 토양에 서식하는 박테리아 Bacillus thuringiensis (Bt)에서 유전자를 분리해 옥수수에 도입해 해충에 강한 옥수수를 만들었습니다. Bacillus thuringiensis는 특정 곤충에게는 독성이 있고 인간, 식물, 또는 다른 동물에겐 무해한 단백질을 생산합니다. 해충에 강한 Bt 옥수수의 도입은 농작물의 수확량을 개선하고 화학 농약의 사용을 줄였습니다. 이러한 농업에서의 응용은 세계적인 식량 공급의 질과 양을 향상시킵니다.

재조합 DNA

개요

과학자들은 실험실에서 서로 다른 출처(종종 다른 종)의 DNA를 결합하여 재조합 DNA를 만듭니다. DNA 클로닝은 ...

Education

교육

JoVE Core

Molecular Biology

분자생물학

Unit 2

제2단원

Studying DNA and RNA

DNA와 RNA 연구

KEY TERMS AND CONCEPTS

주요 용어 및 개념

recombinant dna and gene cloning

Recombinant DNA and gene cloning are tools used to combine DNA from different sources. Scientists use them to isolate a gene, join it to a carrier DNA molecule, and make copies of that DNA.

A plasmid is one common carrier used in this process. A plasmid is a small ring of DNA found in bacteria, and it can carry the inserted gene into a host cell. Once inside the cell, the recombinant DNA can be copied as the cell grows.

This approach is important in biotechnology and genetic research. It helps scientists study genes, produce useful DNA copies, and work with DNA in a controlled way.

Recombinant DNA and Gene Cloning

recombinant dna

Recombinant DNA and gene cloning are tools used to combine DNA from different sources. Scientists use them to isolate a gene, join it to a carrier DNA molecule, and make copies of that DNA. A plasmid is one common carrier used in this process. A plasmid is a small ring of DNA found in bacteria, and it can carry the inserted gene into a host cell. Once inside the cell, the recombinant DNA can be copied as the cell grows. This approach is important in biotechnology and genetic research. It...

dna isolation

Genomic DNA Extraction Methods

Genomic DNA extraction methods separate DNA from proteins, RNA, and other cell material. The process usually follows four main steps. First, the cell structure is disrupted with mechanical or chemical methods to make a cell lysate. Then the DNA is protected from damage during processing. After that, the soluble DNA is separated from cell debris. Finally, the purified DNA is eluted, or washed out, for later use. DNA isolation can follow two main approaches: solution-based methods or solid-phase...

dna agarose gel electrophoresis

Purifying DNA Bands from Agarose Gels

Agarose gel electrophoresis can be used to purify DNA bands after separation by size. In gel extraction, the DNA fragment of interest is cut from the gel and cleaned before later use. This is especially useful in cloning, where pure DNA is needed before ligation, the joining step that links DNA pieces together. In cloning experiments, both the insert and vector DNA fragments are first cut with restriction endonucleases. These fragments often contain enzymes, salts, and other contaminants. Such...

labeling dna probes

DNA Probe Labels for Detecting Targets

DNA probes are short DNA fragments with a reporter tag attached so they can be detected or purified. They bind to matching nucleic acid sequences by complementary base-pairing. That binding helps researchers recover or identify specific regions of DNA. Several reporter tags are commonly used to label DNA probes. These include radioisotopes, fluorophores, and small molecule binding partners such as biotin or digoxigenin. The label choice depends on the probe’s intended use and on the source of...

southern blot

Southern Blot for DNA Detection

Southern blotting is a DNA detection method used after agarose gel electrophoresis separates DNA fragments by size. A DNA ladder with fragments of known length is run beside the sample so the fragment size can be estimated. The blot adds another step when scientists need to confirm the sequence identity of the DNA. First, the target DNA is cut into smaller pieces with a restriction endonuclease and run on an agarose gel. The DNA is then denatured, which means the double strands are separated...

dna microarrays

Gene Chips for Comparing Cell Activity

DNA microarrays are gene chips used to compare gene activity in different conditions. They are high-throughput assays, which means they can analyze large amounts of data at once. They are also relatively inexpensive and can be automated, making them useful for genome-wide studies. These chips are built on glass or silica slides. Probe molecules are covalently attached to the surface through surface functionalization. In many cases, the slides are prepared by chemisorption of silanes to silica...

complementary dna

Making cDNA from RNA Templates

Complementary DNA, or cDNA, is made from an RNA template. It is a DNA copy of RNA, so it can help students study gene expression in a lab setting. cDNA is useful when scientists want to work with genes after they have been turned on in a cell. Because it is copied from RNA, it reflects which genes were active at the time the RNA was collected. This process links RNA and DNA in a simple way. It also gives students a clear view of how genetic information can be copied for analysis.

fish - fluorescent in-situ hybridization

FISH for Chromosome and Pathogen Detection

FISH is a DNA and RNA detection method used in cytogenetics and medical testing. FISH stands for fluorescence in situ hybridization. It uses labeled probes that bind to complementary DNA or RNA sequences on a chromosome or inside a cell. The method was developed in the early 1980s and became widely used because it can show specific genetic targets with high detail. In the past, probes were made by cloning or by reverse transcription of a DNA template. Today, probe oligonucleotides can also be...

pcr - polymerase chain reaction

PCR: DNA Amplification for Genetics Labs

PCR is a DNA amplification method used in genetics labs. PCR stands for polymerase chain reaction. It copies a specific DNA segment so scientists can study small amounts of genetic material. The process uses repeated heating and cooling steps. These steps let the DNA strands separate, primers bind to the target region, and a DNA polymerase build new strands. With each cycle, the target DNA can increase in amount. PCR is a core technique in molecular biology and many school-level biology...

real time rt-pcr

Real-Time RT-PCR for Gene Expression Tracking

Real-time RT-PCR tracks gene expression by measuring how much target DNA is made during PCR. It starts with mRNA, which is converted into complementary DNA, or cDNA, by an enzyme called reverse transcriptase. The cDNA is then amplified by PCR so the target sequence can be measured. This method can be done in one tube as a single-step reaction or in two separate steps. During amplification, the amount of product is followed in real time with fluorescent dyes or sequence-specific probes linked...

race - rapid amplification of cdna ends

Finding cDNA Ends with RACE PCR

Rapid Amplification of cDNA Ends, or RACE, helps scientists capture the full-length cDNA from an mRNA sequence. It works by linking a known internal region to the unknown sequence at either the 5' end or the 3' end. The method is widely used when the missing end of a gene needs to be identified. RACE uses a gene-specific primer that binds to the known cDNA region. A hybrid primer then adds a set anchor sequence to the unknown end of the cDNA. After that, PCR amplifies the DNA segment between...

sanger sequencing

How Sanger Sequencing Reads DNA Fragments

Sanger sequencing reads DNA fragments by separating pieces of different lengths and decoding the order of bases. It is a key method in biology and is often used to study a cloned DNA fragment, a gene mutation, or even much larger genomes. Fredrick Sanger and his collaborators developed the chain-termination method in 1977. The method works by producing a mixture of DNA fragments with variable sizes. These fragments are then separated by capillary gel electrophoresis, which sorts them by length.

next-generation sequencing

DNA Sequencing Methods and Uses

DNA sequencing methods can read millions of DNA fragments at the same time. Next-generation sequencing made this process much faster and much less expensive than older methods. The first human genome project cost $2.7 billion and took 15 years. It was completed in 2003 after major international cooperation among research teams and funding agencies. To prepare DNA for next-generation sequencing, pure genomic DNA is first broken into smaller pieces. These pieces are used to build a sequencing...

rna-seq

RNA-Seq Workflow and Data Analysis

RNA sequencing, or RNA-Seq, is a high-throughput method for studying the transcriptome of a cell. The transcriptome is the full set of RNA molecules in a sample. Transcriptomics helps scientists understand how genomes work, how organisms develop, and how some diseases occur. Before RNA-Seq, researchers often used microarray methods and Sanger sequencing to study RNA. Microarrays had limited coverage and depended on already knowing the genome sequence. Sanger sequencing was low-throughput,...

genome annotation and assembly

How Genome Assemblies Are Updated

Genome assembly and genome annotation are key steps in genome analysis. Genome assembly puts DNA sequencing data back in the correct order to make a close copy of the original genome. Genome annotation then identifies functional parts on that assembled genome. Assembling a genome is not simple. In human populations, gene copy number can vary, and repeated DNA sequences can make the process harder. Even so, the physical location of human genes stays constant. Bacterial genomes can be more...

restriction enzymes

DNA Cutting Sites and Bacterial Defense

Restriction enzymes are bacterial proteins that cut DNA at specific sequence sites. They recognize palindromic DNA sequences, also called inverted repeats, and cut the DNA in a sequence-specific way. These sites are often found in regions with important functions, such as the origin of replication, gene operator sites, and transcription termination signals. Bacteria protect their own DNA by methylating these same sites. Methylation means adding chemical marks that keep the restriction enzyme...

maxam-gilbert sequencing

Chemical Cleavage DNA Sequencing

Chemical cleavage DNA sequencing uses specific chemicals to cut DNA at known sites. Allan Maxam and Walter Gilbert described this method in the same year that the Sanger sequencing method was discovered. It reads the DNA sequence by breaking purified DNA into fragments and then studying the fragment pattern. The method depends on chemicals that cleave DNA at specific bases. The resulting fragments are separated by electrophoresis, which is a way to sort DNA pieces by size using an electric...

key terms and concepts

Learn how recombinant DNA and gene cloning work by joining a gene to a plasmid and copying it in a host cell for biotechnology and research.

Recombinant DNA and Gene Cloning | JoVE K12

molecular biology

JoVE Core 분자생물학은 명확하고 간결한 애니메이션을 통해 분자 단계에서 일어나는 세포 과정을 설명합니다.또한, 과학자가 직접 실험을 진행하는 실사 영상을 통해 분자생물학 개념의 응용 사례를 보여줍니다.

Virginia  Standards of Learning

HIgh School

BIO.5

BIO.5.e

Alabama Course of Study Science

9-12

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

From Molecules to Organisms: Structures and Processes

9-12.B.1.2.c

Florida Department of Education

Life Science

SC.912.L.16

SC.912.L.16.12

Texas Essential Knowledge and Skills

BIO.7

BIO.7.D

myJoVE Corporation

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In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled ...

Obtain, evaluate, and communicate information on how molecular biotechnology may be used in a variety of fields.
EXPLANATION: The goal is to help students appreciate that the basic science knowledge being studied can be manipulated and used to design useful tools for further scientific investigations, medical treatments, agricultural yields, etc. A detailed understanding of any technical procedures is not expected, but rather a "big picture" view of concepts the technology utilizes and the applications the technology is being used for. These technologies may be best investigated along with the standards from LS1 From Molecules to Organisms: Structure and Functions, as the students are learning about DNA and protein. Comparative DNA and protein sequence analysis is also a large part of current phylogenic research and could accompany LS4 Biological Change: Unity and Diversity. Students may also consider the pros and cons of biotechnical applications. Investigation of techniques utilized could come from field trips to witness the use of these techniques in practice, lab exercises, virtual labs, simulation activities, interviews with professionals, etc. Molecular techniques could include: PCR, electrophoresis, restriction enzyme digestion of DNA, DNA sequencing, plasmid-based transformation, transfection, etc. These techniques are used in fields of medicine, agriculture, biomedical engineering, forensic science, etc.

Karyotyping

Lampbrush Chromosomes

In 1882, Flemming observed lampbrush chromosomes (LBC) in salamander eggs. Later in 1892, R&#252;ckert observed LBCs in shark egg cells and coined the ...

Polytene Chromosomes

Polytene chromosomes are giant interphase chromosomes with several DNA strands placed side by side. They were discovered in the year 1881 by Balbiani in ...

Euchromatin

The extent of chromatin compaction can be studied by staining chromatin using specific DNA binding dyes. Under the microscope, the dense-compacted regions ...

Heterochromatin

X and Y Chromosomes

Among mammals, the gender of an organism is determined by the sex chromosomes. Humans have two sex chromosomes, X and Y. Every human diploid cell has 22 ...

The Y Chromosome Determines Maleness

The Y chromosome is a sex chromosome&#160;found in several vertebrates and mammals, including humans. In addition to 22 pairs of autosomes, the human ...

The Ratio of X Chromosome to Autosomes

In most organisms, sex is determined by the ratio of X and Y chromosomes. However, in some organisms, such as Drosophila and C.elegans, sex is determined ...

Investigate means by which karyotypes are utilized in diagnostic medicine.
EXPLANATION: Engineers incorporate growing scientific knowledge in order to increase benefits to humans and decrease cost and risks. Karyotyping technology and its use in medical diagnosis can complement the standards from LS3: Hereditary: Inheritance, and Variation of Traits, as the students are learning about chromosomal organization of genomic information that determines one's phenotype. Analyze a large set of karyotypes to identify common patterns. Identify variations from the most commonly observed patterns and correlate them with patient phenotypes to propose a cause and effect relationship, discussing limitations of correlative data interpretation.

BIO1.ETS2.2

Whole Body Regeneration

Regeneration is the process of restoring injured or lost tissues, organs, or&#160;body parts. While simpler organisms generally show greater ability to ...

Analyze scientific and ethical arguments to support the pros and cons of applications of a specific biotechnology technique such as stem cell usage, in vitro fertilization, or genetically modified organisms.
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.

BIO1.ETS2.3

Recombinant DNA and Gene Cloning

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