Name: recombinant dna in cloning and gmos
Uploaded: 2026-08-26T08:20:51.528Z
Duration: 1 min 34 s
Channel: JoVE

- Il DNA ricombinante &#xE8; DNA

da fonti differenti, di
solito da specie differenti,

che si combina insieme nel laboratorio

per ampi usi nella ricerca
clinica e scientifica.

Il DNA ricombinante consiste
di un gene di interesse,

qui l&#x27;insulina da un organismo donatore

inseriti in un vettore,

DNA autoreplicamente
da un altro organismo,

come un virus o un plasmide,
un piccolo pezzo circolare

di DNA batterico.

Il DNA da ciascuna risorsa
&#xE8; per prima cosa tagliato

da endonucleasi di restrizione,

enzimi che tagliano il DNA
in una sequenza precisa.

Il DNA del donatore &#xE8; collegato
a quello del DNA vettore,

con l&#x27;aiuto di enzimi DNA ligasi.

Il DNA ricombinante si inserisce
in un organismo ospite,

batteri o lievito, dove si replica,

creando molte copie del gene di interesse.

Questo gene pu&#xF2; essere
studiato pi&#xF9; facilmente

o, come con l&#x27;insulina,

trasformato in proteina usando
il meccanismo cellulare.

Khan, Suliman, Muhammad Wajid Ullah, Rabeea Siddique, Ghulam Nabi, Sehrish Manan, Muhammad Yousaf e Hongwei Hou. 2016. "Ruolo della tecnologia del DNA ricombinante per migliorare la vita". Giornale internazionale di genomica 2016. [<a href="https://doi.org/10.1155/2016/2405954" target="__blank">Sorgente</a>]

Matsumura, Ichiro. 2015. "Perché Johnny non può clonare: insidie comuni e soluzioni non così comuni". BioTechniques 59 (3): IV–XIII. [<a href="https://doi.org/10.2144/000114324" target="__blank">Sorgente</a>]

dna ricombinante

Recombinant DNA Technology

<h4>Panoramica</h4>

Gli scienziati creano DNA ricombinante combinando il DNA di fonti diverse, spesso altre specie, in laboratorio. La clonazione del DNA consente ai ricercatori di studiare geni specifici inserendoli in cellule facilmente manipolate, come i batteri. Gli organismi che contengono DNA ricombinante sono noti come organismi geneticamente modificati (OGM). La tecnologia del DNA ricombinante produce organismi con nuovi geni che possono beneficiare la scienza, la medicina e l&#x27;agricoltura.

<h4>Come fanno gli scienziati a creare DNA ricombinante?</h4>

La creazione di DNA ricombinante comporta l&#x27;inserimento di un gene di interesse in un vettore, un veicolo che trasporta il DNA estraneo nelle cellule ospiti per la replicazione del DNA e l&#x27;espressione proteica. I vettori di clonazione pi&#xF9; comunemente utilizzati sono plasmidi, piccoli pezzi circolari di DNA che si replicano indipendentemente dal DNA cromosomico dell&#x27;ospite.

Per creare DNA ricombinante, sia il DNA del donatore, compreso il gene di interesse, sia il vettore vengono tagliati in specifiche sequenze di nucleotidi, chiamate siti di restrizione, utilizzando enzimi di restrizione. L&#x27;enzima DNA ligase sigilla la spina dorsale zucchero-fosfato dove si connettono il gene di interesse e il plasmide.

Il risultato &#xE8; una molecola di DNA ricombinante costituita da un vettore con un pezzo integrato di DNA del donatore, chiamato inserto. Uno scienziato pu&#xF2; quindi introdurre questa molecola di DNA ibrido in un organismo ospite, tipicamente batteri o lieviti, dove si replica facilmente e rapidamente. Questo crea molte copie del gene di interesse, che &#xE8; necessario per la ricerca scientifica e altre applicazioni. Il gene pu&#xF2; anche essere trascritto e tradotto nella proteina desiderata, come l&#x27;insulina umana, utilizzando il meccanismo cellulare dell&#x27;ospite.

La creazione di DNA ricombinante &#xE8; un processo imperfetto e spesso si verificano errori. Ad esempio, il vettore potrebbe chiudersi senza l&#x27;inserto o l&#x27;inserto potrebbe non essere corretto (ad esempio, all&#x27;indietro). Prima di utilizzare il DNA ricombinante per ulteriori studi, i ricercatori devono verificare la presenza di errori. Il sequenziamento dei nucleotidi pu&#xF2; aiutare a identificare le colonie di batteri che trasportano plasmidi con l&#x27;inserto corretto.

Gli scienziati utilizzano il DNA ricombinante per studiare geni e proteine

La tecnologia del DNA ricombinante &#xE8; particolarmente vantaggiosa quando uno scienziato ha bisogno di molte copie di un gene di interesse o di un prodotto proteico. Tuttavia, la ricerca di uno scienziato pu&#xF2; richiedere un ulteriore livello di complessit&#xE0;, come il rilevamento o la purificazione della proteina desiderata. Per raggiungere questo obiettivo, un ricercatore pu&#xF2; allegare un tag o un reporter, proteine utilizzate per identificare un prodotto genetico, alla proteina desiderata per creare un gene di fusione, o gene chimerico.

<h4>Applicazioni in Medicina e Agricoltura</h4>

Gli scienziati hanno usato per la prima volta la tecnologia del DNA ricombinante per produrre insulina umana nei batteri, con conseguente trattamento per il diabete. Da quella scoperta iniziale, i ricercatori hanno generato altre DLL ricombinanti per uso terapeutico. I batteri ricombinanti rendono l&#x27;ormone della crescita umano, una proteina necessaria per la normale crescita e sviluppo, per trattare i pazienti con carenza di ormone della crescita. Le cellule di mammiferi ricombinanti, derivate da esseri umani e criceti, producono il fattore VIII, una proteina necessaria per la normale coagulazione del sangue, per il trattamento di pazienti con emofilia. Evidentemente, la tecnologia del DNA ricombinante &#xE8; un potente strumento per la produzione su larga scala di proteine essenziali.

I progressi agricoli nella tecnologia del DNA ricombinante influenzano anche il benessere umano. Ad esempio, gli allevatori di mais hanno subito notevoli danni alle colture a causa del borer di mais europeo. In risposta, gli scienziati hanno isolato i geni da un batterio che abita il suolo (Bacillus thuringiensis (Bt) - per creare mais geneticamente modificato e resistente ai parassiti. Bacillus thuringiensis produce naturalmente proteine tossiche per alcuni insetti, ma non per gli esseri umani, le piante o altri animali. L&#x27;introduzione del mais Bt resistente ai parassiti ha migliorato le rese delle colture e ridotto l&#x27;uso di pesticidi chimici. Tali applicazioni agricole migliorano la qualit&#xE0; e la quantit&#xE0; dell&#x27;approvvigionamento alimentare globale.

DNA ricombinante

Overview

Scientists create recombinant DNA by combining DNA from different sources&#x2014;often, other species&#x2014;in the laboratory. DNA cloning allows ...

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recombinant dna in cloning and gmos

Recombinant DNA is made by joining DNA from different sources in the lab. Scientists often use it to clone genes and study them in cells that are easy to handle, such as bacteria. Organisms that carry recombinant DNA are called genetically modified organisms, or GMOs. This technology gives scientists new tools for research, medicine, and agriculture.

To build recombinant DNA, researchers place a gene of interest into a vector, which is a DNA carrier that moves foreign DNA into a host cell. The most common vectors are plasmids, small circular DNA molecules that copy themselves separately from the cell’s chromosomal DNA. Scientists cut both the donor DNA and the vector at specific restriction sites using restriction enzymes. Then DNA ligase seals the sugar-phosphate backbone where the insert joins the plasmid.

The final product is a recombinant DNA molecule with a vector carrying an inserted piece of donor DNA. Scientists can put this hybrid DNA into a host organism, usually bacteria or yeast. The host then replicates the DNA quickly, making many copies of the gene of interest. In some cases, the host’s machinery also transcribes and translates the gene into a protein, such as human insulin.

Creating recombinant DNA is not always perfect. The vector may close without taking in the insert, or the insert may end up in the wrong direction. Before moving on, researchers check for these errors. Nucleotide sequencing helps them find bacteria colonies that contain plasmids with the correct insert.

Recombinant DNA is useful when scientists need many copies of a gene or a protein product. Some projects need extra steps, such as making the protein easier to detect or purify. For that reason, researchers may attach a tag or reporter protein to create a fusion gene, also called a chimeric gene.

This technology has major uses in medicine and agriculture. Scientists first used recombinant DNA to make human insulin in bacteria for diabetes treatment. They also use recombinant bacteria to produce human growth hormone for growth hormone deficiency. Recombinant mammalian cells, from humans and hamsters, make Factor VIII to treat hemophilia. In agriculture, scientists used genes from Bacillus thuringiensis, or Bt, to create pest-resistant corn. Bt proteins are toxic to certain insects, not humans, plants, or other animals, so Bt corn can improve crop yields and reduce chemical pesticide use.

Recombinant DNA in Cloning and GMOs

what is genetic engineering?

How Scientists Edit DNA for New Traits

Genetic engineering changes an organism’s DNA to add useful traits. Scientists use it in bacteria, plants, animals, and even viruses for research, medicine, agriculture, and industry. It can bring major benefits, but it also raises ethical concerns about humans and the food supply. This work is possible because DNA has the same basic structure and genetic code across life. That shared code lets scientists modify DNA in several ways. One method is CRISPR/Cas9, often called molecular scissors,...

dna isolation

Steps in DNA Extraction

DNA extraction is the process of removing and purifying DNA from cells. It is used in many biotechnology and research applications, including molecular cloning. The main goal is to free DNA from the cell and prepare it for later study or use. The first step is cell lysis, which means breaking cells open. Researchers may use a homogenizer, sonicator, or bead beater to physically lyse the cells. They also often add detergent to chemically disrupt the lipid-based cell membranes. This helps...

recombinant dna technology

Recombinant DNA is made by joining DNA from different sources in the lab. Scientists often use it to clone genes and study them in cells that are easy to handle, such as bacteria. Organisms that carry recombinant DNA are called genetically modified organisms, or GMOs. This technology gives scientists new tools for research, medicine, and agriculture. To build recombinant DNA, researchers place a gene of interest into a vector, which is a DNA carrier that moves foreign DNA into a host cell. The...

PCR Cycle for DNA Amplification

PCR is a laboratory method for making many copies of a DNA segment. It can turn a small amount of DNA into millions or even billions of copies in just a few hours. The process uses a heat-resistant DNA polymerase inside a machine called a thermocycler. PCR was developed by Kary Mullis in 1983, and he received the 1993 Nobel Prize in Chemistry for it. The method is fast, relatively inexpensive, and precise. It has become useful for molecular cloning, gene mutagenesis, pathogen detection, gene...

complementary dna

From mRNA to cDNA for Gene Studies

Complementary DNA, or cDNA, is made from messenger RNA (mRNA) so scientists can study which genes are active in cells and tissues. Only genes that are transcribed into mRNA are expressed. By extracting mRNA from cells, researchers can compare gene expression across samples. cDNA is produced by reverse transcription. This process copies mRNA into DNA, which is useful because mRNA does not contain introns, or non-coding regions, and other regulatory sequences. As a result, cDNA can help...

antibiotic selection

Antibiotic Resistance in Cloning

Antibiotic resistance genes help scientists find bacteria that took up a plasmid with a gene of interest. In genetic engineering, this resistance is used as a marker, not just as a medical concern. The same process also connects to the growing problem of resistant bacteria in the real world. Antibiotics, such as penicillin, stop bacterial growth or kill bacteria. When bacteria naturally have, or are engineered to carry, an antibiotic resistance gene, the antibiotic no longer works on them.

transgenic organisms

Creating Transgenic Plants and Animals

Transgenic organisms are genetically engineered to carry a transgene, which is a gene from a different species, in their genome. The inserted gene may be a new gene for that organism or a different version of one it already has. Scientists usually make transgenes with recombinant DNA and DNA cloning methods. Researchers use transgenic bacteria, plants, and animals to answer biological questions and to build useful tools. To make a transgenic organism, they first choose a transgenesis method,...

in-vitro mutagenesis (gene knockouts)

Knockout Mice and Gene Replacement

Knockout mice help researchers study gene function by showing what happens when a gene is turned off. These engineered animals are also useful for modeling human diseases such as cancer, Parkinson’s disease, and diabetes. A specific gene can be targeted by using a targeting vector, which is a piece of engineered DNA that replaces the normal gene and inactivates it. The vector has matching DNA sequences on both ends. These homologous sequences let the vector swap in for the target gene through...

crispr

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...

gene therapy

Gene Delivery for Serious Diseases

Gene therapy is a technique used to treat serious disease by adding a gene to a person’s cells. The added gene may be a healthy copy of a mutated gene. It may also be a different gene that inactivates or compensates for the disease-causing gene. One example is severe combined immunodeficiency, or SCID, caused by a mutation in the gene for adenosine deaminase. In that case, a working copy of the gene can be inserted so the patient’s cells make the enzyme again. This can cure the disease in some...

embryonic stem cells

How Embryonic Stem Cells Become Specialized

Embryonic stem cells are pluripotent cells that have not yet specialized. Pluripotent means they can give rise to many different cell types in the body. Because of this, they are useful in science and medicine for making cells needed in research or for replacing cells lost to damage or disease. These cells come from the inner cell mass of a blastocyst, which is an early embryo about 3 to 5 days after fertilization in humans. At this stage, the embryo has not yet implanted in the uterus. Human...

adult stem cells

Adult Stem Cells in the Small Intestine

Adult stem cells help maintain tissues by replacing old or damaged cells. These undifferentiated cells divide to make more stem cells or progenitor cells, which then become mature, specialized cells. Small groups of adult stem cells are found in many tissues, including the bone marrow, brain, skin, and gut. The small intestine shows this process clearly. Its lining, called the epithelium, is renewed continuously by adult stem cells. It is also the fastest replaced tissue in the human body,...

induced pluripotent stem cells

Reprogramming Mature Cells into Stem Cells

Reprogramming mature cells into stem cells lets scientists turn specialized cells back into an unspecialized state. These cells are called induced pluripotent stem cells, or iPSCs. Like embryonic stem cells, iPSCs are pluripotent, which means they can form all cell types. To make iPSCs, researchers start with mature cells such as skin fibroblasts or blood cells. The cells are grown in culture, and genes for several transcription factors are delivered with a viral vector. The cells then make...

reproductive cloning

Cloning Animals with SCNT

Reproductive cloning can produce a genetically identical copy of an entire organism. In adult animals, this is usually done with somatic cell nuclear transfer, or SCNT. Another way clones can form is by splitting an early embryo, which is similar to how identical twins arise naturally. In SCNT, an egg cell is taken from an animal and its nucleus is removed. This creates an enucleated egg. A somatic cell, which is any body cell that is not a sex cell, is then taken from the animal to be cloned.

genomics

Genomics and DNA Sequencing

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

See how scientists build recombinant DNA with plasmids, restriction enzymes, and ligase, then use it to make proteins and GMOs in medicine and farming.

Recombinant DNA in Cloning and GMOs | JoVE K12

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Recombinant DNA in Cloning and GMOs

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