0
Why is it important to use the same restriction enzyme for both cells in recombinant DNA?
Wiki User
∙ 9y ago
Restriction enzymes are endonucleases that digest the DNA at a sequence specific site. Hind III for example cut between two As in the sequence AAGCTT in the both strand forming a sticky end. If you use this enzyme to cut in your vector DNA, you have to use the same enzyme in the insert DNA so as they can ligate by DNA ligation. This is the important use of same restriction enzyme in cloning.
Wiki User
∙ 9y ago
Add your answer:
Earn +20 pts
What is a basic technique for gene transfer?
Firstly, the plasmid is removed from the bacteria where it is cut open by restriction enzymes. The desired DNA is then extracted from the donor, which is then cut open with the same restriction enzyme. This results in sticky ends and the two pieces of DNA from the two organisms can hence be mixed with complementary base sequences. Ligase is then used to splice the pieces of DNA to form recombinant plasmids, which is then inserted into host cells.
How did Boyer and Cohen performe their recombinant DNA experiment?
Herbert Boyer and Stanley Cohen created the first recombinant DNA organism using recombinant DNA technology, or gene splicing, which allows the manipulation of DNA. They showed that the gene for a frog ribosomal RNA could be transferred and expressed in bacterial cells. Boyer and Cohen removed plasmids, small rings of DNA located in a cell's cytoplasm, from a cell. Using restriction enzymes, they cut the DNA at precise positions and then recombined the DNA strands in their own way using DNA ligase enzyme. They then inserted the altered DNA into E. coli bacteria. The bacterial cells could be made to produce specific proteins using gene splicing. This technology was a major breakthrough for genetic engineering. Their experiments dramatically demonstrated the potential impact of DNA recombinant engineering on medicine and pharmacology, industry and agriculture.
What recombinant cells can correctly attach sugars to proteins to form glycoprotein products?
Mammalian Cells
How does bacterial cell protect its own DNA from restriction enzymes?
In order to protect the bacterial genomic DNA from its own restriction enzymes, bacterial cells employ a system, wherein methyl transferases methylate certain bases on the DNA sequence, making them unrecognizable to the restriction enzymes.Each restriction enzyme has a methylase associated with it on the chromosome. This methylase puts methyl groups on the host DNA, and the restriction enzyme doesn't recognize its recognition sequence when it is so methlyated. The host DNA is thus protected from the actions of its own restriction enzyme.Incoming (foreign) DNA is unlikely to be protected (methylated) in the same manner, thus this invading DNA is digested by the hosts restriction enzyme(s).When working in cloning experiments, the principle is the same -- DNA to be digested is carried by a plasmid in a host that does not methylate DNA in the pattern that would cause the restriction enzyme to see it as protected, thus it is cut. DNA generated by PCR is similarly unmethylated, and is therefore also digested.Some enzymes won't cut DNA isolated from dam+ or dcm+ hosts (two common bacterial methylases), thus one must know the genotype of the host cloning strain if using a restriction enzyme whose action is blocked by dam ordcmmethylation.
What are some current recombinant technologies?
Production of human insulin from bacterial cells.
Why restriction enzyme cannot cut its own DNA?
Restriction enzymes are produced by bacteria to help destroy foreign, invading DNA, such as the DNA of bacteriophage (a virus that infects bacterial cells). Every restriction enzyme comes with a methylase enzyme, or more specifically, a DNA methyltransferase. The methylase enzyme methylates (adds a methyl group) to the restriction endonuclease site on the cell's own DNA, which protects the sites from the restriction enzyme so that it does not degrade its own DNA.
What is a basic technique for gene transfer?
Firstly, the plasmid is removed from the bacteria where it is cut open by restriction enzymes. The desired DNA is then extracted from the donor, which is then cut open with the same restriction enzyme. This results in sticky ends and the two pieces of DNA from the two organisms can hence be mixed with complementary base sequences. Ligase is then used to splice the pieces of DNA to form recombinant plasmids, which is then inserted into host cells.
What enzyme is used to dissolve heart tissue into individual cells?
First a specialized detergent is used without affecting the integrity of the protein in the tissue and then recombinant endonuclease is used to degrade Nucleic acid.
How did Boyer and Cohen performe their recombinant DNA experiment?
Herbert Boyer and Stanley Cohen created the first recombinant DNA organism using recombinant DNA technology, or gene splicing, which allows the manipulation of DNA. They showed that the gene for a frog ribosomal RNA could be transferred and expressed in bacterial cells. Boyer and Cohen removed plasmids, small rings of DNA located in a cell's cytoplasm, from a cell. Using restriction enzymes, they cut the DNA at precise positions and then recombined the DNA strands in their own way using DNA ligase enzyme. They then inserted the altered DNA into E. coli bacteria. The bacterial cells could be made to produce specific proteins using gene splicing. This technology was a major breakthrough for genetic engineering. Their experiments dramatically demonstrated the potential impact of DNA recombinant engineering on medicine and pharmacology, industry and agriculture.
What recombinant cells can correctly attach sugars to proteins to form glycoprotein products?
Mammalian Cells
How could you use genetic engineering techniques to make transformed bacteria that produce the enzyme?
Extract DNA from the cells of people who can make the digestion enzyme. Cut the DNA with restriction enzymes to cut out the gene that codes for the enzyme. Use gel electrophoresis to locate the gene. Then, use polymerase chain reaction to make copies of the gene. Choose a plasmid that has an antibiotic-resistance genetic marker, and cut the plasmid with the smae restriction enzyme use to cut out the hyman gene. Insert the copies of the human gene into the plasmids. Allow bacterial cells to take in the plasmids. Select for transformed bacteria by growing them in a culture containing the antibiotic. These bacteria will make the digestion enzyme.
What are some current recombinant technologies?
Production of human insulin from bacterial cells.
How does bacterial cell protect its own DNA from restriction enzymes?
In order to protect the bacterial genomic DNA from its own restriction enzymes, bacterial cells employ a system, wherein methyl transferases methylate certain bases on the DNA sequence, making them unrecognizable to the restriction enzymes.Each restriction enzyme has a methylase associated with it on the chromosome. This methylase puts methyl groups on the host DNA, and the restriction enzyme doesn't recognize its recognition sequence when it is so methlyated. The host DNA is thus protected from the actions of its own restriction enzyme.Incoming (foreign) DNA is unlikely to be protected (methylated) in the same manner, thus this invading DNA is digested by the hosts restriction enzyme(s).When working in cloning experiments, the principle is the same -- DNA to be digested is carried by a plasmid in a host that does not methylate DNA in the pattern that would cause the restriction enzyme to see it as protected, thus it is cut. DNA generated by PCR is similarly unmethylated, and is therefore also digested.Some enzymes won't cut DNA isolated from dam+ or dcm+ hosts (two common bacterial methylases), thus one must know the genotype of the host cloning strain if using a restriction enzyme whose action is blocked by dam ordcmmethylation.
What is the role of enzyme in cells?
enzyme sped up the chemical reaction
Are plasmid vectors helpful?
Plasmid vectors are an invaluable genetic engineering tool for inserting recombinant DNA sequences into different organisms or cells in culture.Plasmids are essentially circular DNA constructs composed of some essential elements like:An origin of replicationA multiple cloning site which consists of restriction sites where the recombinant DNA can be insertedMarker genes (like antibiotic resistance)reporter genes to confirm a successful transformation
How can a bacterium produce restriction enzymes that do not cleave its DNA?
In order to protect the bacterial genomic DNA from its own restriction enzymes, bacterial cells employ a system, wherein methyl transferases methylate certain bases on the DNA sequence, making them unrecognizable to the restriction enzymes.Each restriction enzyme has a methylase associated with it on the chromosome. This methylase puts methyl groups on the host DNA, and the restriction enzyme doesn't recognize its recognition sequence when it is so methlyated. The host DNA is thus protected from the actions of its own restriction enzyme.Incoming (foreign) DNA is unlikely to be protected (methylated) in the same manner, thus this invading DNA is digested by the hosts restriction enzyme(s).When working in cloning experiments, the principle is the same -- DNA to be digested is carried by a plasmid in a host that does not methylate DNA in the pattern that would cause the restriction enzyme to see it as protected, thus it is cut. DNA generated by PCR is similarly unmethylated, and is therefore also digested.Some enzymes won't cut DNA isolated from dam+ or dcm+ hosts (two common bacterial methylases), thus one must know the genotype of the host cloning strain if using a restriction enzyme whose action is blocked by dam ordcmmethylation.
Why is fermentation an important process in cells?
Fermentation is the breakdown of carbohydrates by enzyme action under anaerobic conditions. This is important to provide energy when insufficient oxygen is present.
