0
Why is it important to use the same restriction enzyme for both cells in recombinant DNA?
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.
What else can I help you with?
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.
How will you identify cells carrying the recombinant plasmid?
The transformants are selected for on agar containing an appropriate antibiotic. For example if your recombinant plasmid contains a kanamycin cassette, then only the cells containing the plasmid will grow on an agar plate containing kanamycin. PCR can then be performed on the colonies to ensure they contain your gene of interest on the plasmid.
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 is a collection of cells containing DNA fragments produced by restriction enzymes and incorporated into plasmids called?
A DNA LibraryA collection of cells containing DNA fragments produced by restriction enzymes and incorporated into plasmids is called a DNA library. RNA can manufacture DNA via the action of reverse transcriptase.
How does a human insulin genes become part of a plasmid?
1. Scientists remove plasmids, small rings of DNA, from bacterial cells. 2. An enzyme cuts open the plasmid DNA. The same enzyme removes the human insulin gene from its chromosome. 3. The human insulin gene attaches the open ends of the plasmid to form a closed ring. 4. Some bacterial cells take up the plasmids that have the insulin gene. 5. When cells reproduce, the news cells will contain copies of the engineered plasmid. The foreign gene directs the cell to produce human insulin.
What is the best explanation for why a restriction enzyme does not cut the DNA of the cell that produces it?
Restriction enzymes are made by cells to protect their own DNA from being cut. These cells produce a modification enzyme that adds a methyl group to specific sites on their own DNA sequence, which prevents the restriction enzyme from cutting. This process is known as "methylation protection."
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.
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.
How will you identify cells carrying the recombinant plasmid?
The transformants are selected for on agar containing an appropriate antibiotic. For example if your recombinant plasmid contains a kanamycin cassette, then only the cells containing the plasmid will grow on an agar plate containing kanamycin. PCR can then be performed on the colonies to ensure they contain your gene of interest on the plasmid.
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 are the requirements for recombinant DNA technology?
Requirements for recombinant DNA technology include a vector (such as a plasmid or virus) to carry the desired DNA fragment, restriction enzymes to cut the DNA at specific sites, and DNA ligase to join the DNA fragments together. Additionally, cells capable of taking up and expressing the recombinant DNA are needed, along with appropriate selection markers to identify successfully transformed 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.
What is a collection of cells containing DNA fragments produced by restriction enzymes and incorporated into plasmids called?
A DNA LibraryA collection of cells containing DNA fragments produced by restriction enzymes and incorporated into plasmids is called a DNA library. RNA can manufacture DNA via the action of reverse transcriptase.
How does a human insulin genes become part of a plasmid?
1. Scientists remove plasmids, small rings of DNA, from bacterial cells. 2. An enzyme cuts open the plasmid DNA. The same enzyme removes the human insulin gene from its chromosome. 3. The human insulin gene attaches the open ends of the plasmid to form a closed ring. 4. Some bacterial cells take up the plasmids that have the insulin gene. 5. When cells reproduce, the news cells will contain copies of the engineered plasmid. The foreign gene directs the cell to produce human insulin.
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.
How is recombinant replicated?
Recombinant DNA is replicated using host cells, typically bacteria or yeast, that have been engineered to contain the desired DNA sequence. These host cells are then grown in a lab setting under specific conditions that allow for the replication of the recombinant DNA. The cell division process allows for the production of multiple copies of the recombinant DNA.
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.
