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Why is a restriction enzyme that cuts your plasmid more than once unusable?
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∙ 11y ago
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∙ 11y ago
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Which Restriction enzyme are studied in Recombinant DNA Technology?
It's not the restriction enzymes that are studied, its the DNA. The enzyme cuts or "restricts" the DNA strand at a known sequence of nucleotides. Different enzyme, different sequence. For a Biomanufacturing application, where we want to insert foreign DNA, the gene of interest is cut and spliced with a restriction enzyme into a recombinant plasmid, transformed into a bacteria, and sent merrily on it's way to make Insulin, or whatever. With an unknown piece of DNA (a functional gene that makes a protein of interest or is being studied), the plasmid has "restriction sites" or nucleotide sequences, for several restriction enzymes, all of which I have mapped out. The unknown piece of DNA is cut at each end by a single restriction enzyme and inserted into the plasmid, which gives me some landmarks. I insert the plasmid into a bacteria, grow a culture so the bacteria makes many millions of copies of the plasmid, extract the plasmid, and run an experiment called a restriction digest. The restriction digests are a series of reaction with single enzyme and combinations of two and three enzymes, all cutting the plasmid at different nucleotide sequences. Then I run an agarose gel electrophoresis, which separates all the different pieces of DNA by size, and do an analysis called a Restriction Map. This counts the DNA fragments and their sizes, which enzyme and combination of enzymes produced which sizes and how many fragments, which enzyme cuts where, which cuts were definitely in the known part of the plasmid, which were probably in the unknown DNA, adding up nucleotide sequence numbers to make sure different mapping guesses agree, etcetera, etcetera, and so forth. Until at last, a map of the size and restriction sites of the unknown DNA insert into the known plasmid vector is deduced. This used to be done by hand, but there are computer programs that do it now. This is Research, the Technology is down the line a few steps when the gene has been characterized, the protein produced has been characterized, the trials are done, and the restriction enzyme to insert the gene into the bacteria for Bioman has been established
What type of the enzyme that cuts the bonds of DNA molecule at the origin of replication?
restriction endonucleases
What is the restriction site of the restriction enzyme Hae III?
The restriction site of Hae III is GGCC. It cuts between the G and the C. This produces blunt ends.
What cuts bacterial plasmids?
Enzymes called restriction endonucleases can cut plasmids. However, in order for a cut to be produced, the plasmid should contain a specific sequence of nucleotides called the restriction site
What are restriction enzymes Explain the significance of these enzymes in recombinant DNA technology.?
A restriction enzyme (or restriction endonuclease) is an enzyme that cuts double-stranded or single stranded DNA at specific recognition nucleotide sequences known asrestriction sites....................refer in this website en.wikipedia.org/wiki/Restriction_enzyme
An enzyme that cuts double-stranded DNA at specific nucleotide sequences?
Such an enzyme is called a restriction endonuclease
Which Restriction enzyme are studied in Recombinant DNA Technology?
It's not the restriction enzymes that are studied, its the DNA. The enzyme cuts or "restricts" the DNA strand at a known sequence of nucleotides. Different enzyme, different sequence. For a Biomanufacturing application, where we want to insert foreign DNA, the gene of interest is cut and spliced with a restriction enzyme into a recombinant plasmid, transformed into a bacteria, and sent merrily on it's way to make Insulin, or whatever. With an unknown piece of DNA (a functional gene that makes a protein of interest or is being studied), the plasmid has "restriction sites" or nucleotide sequences, for several restriction enzymes, all of which I have mapped out. The unknown piece of DNA is cut at each end by a single restriction enzyme and inserted into the plasmid, which gives me some landmarks. I insert the plasmid into a bacteria, grow a culture so the bacteria makes many millions of copies of the plasmid, extract the plasmid, and run an experiment called a restriction digest. The restriction digests are a series of reaction with single enzyme and combinations of two and three enzymes, all cutting the plasmid at different nucleotide sequences. Then I run an agarose gel electrophoresis, which separates all the different pieces of DNA by size, and do an analysis called a Restriction Map. This counts the DNA fragments and their sizes, which enzyme and combination of enzymes produced which sizes and how many fragments, which enzyme cuts where, which cuts were definitely in the known part of the plasmid, which were probably in the unknown DNA, adding up nucleotide sequence numbers to make sure different mapping guesses agree, etcetera, etcetera, and so forth. Until at last, a map of the size and restriction sites of the unknown DNA insert into the known plasmid vector is deduced. This used to be done by hand, but there are computer programs that do it now. This is Research, the Technology is down the line a few steps when the gene has been characterized, the protein produced has been characterized, the trials are done, and the restriction enzyme to insert the gene into the bacteria for Bioman has been established
Hi does any one know if I want to put some restriction enzyme close to cDNA should I put some nucleotide befor this restriction enzyme?
Restriction enzyme cuts DNA strand at specific locations Restriction enzyme cuts DNA strand at specific locations
Based on restriction maps of plasmid determine the number of DNA fragments and sizes of the fragments?
Plasmids are circular pieces of DNA, so the number of fragments equals the number of cuts from the restriction enzymes. You can easily see this if you start with one restriction enzyme that cuts the plasmid in only one place. Cutting the circle in one place yields you only one fragment. If the restriction cuts in two places, you end up with two fragments; with three places, three fragments, etc. With linear chromosomes, the situation is different. Cutting a linear chromosome in one place yields two fragments, cutting in two places yields three fragments, etc. So the number of fragments is always one more than the number of cuts. A restriction map of a plasmid will show all of the cuts the restriction enzymes made. Each cut is labeled with the enzyme that made it. One can count the spaces between cuts to determine the number of fragments that are produced. Restriction maps usually (but not always) also show the size of each fragment.
What is the term that describes the sequence of DNA that a restriction enzyme finds and cuts?
Template Sequence
Why must you use an enzyme that will not cut anywhere within the gene that you are inserting into a plasmid?
If you are trying to take a gene from a DNA strand and put insert it into a plasmid, you wouldn't want a restriction enzyme to cut that gene up, or else it would be pretty useless. In other words, you need an enzyme or two that cuts outside that gene so that it can be functional after it's inserted into a plasmid. After your gene of interest is inserted into a plasmid, the plasmid can be put back into a bacterium, then you could genetically engineer plants with it or let the bacterium reproduce and produce many copies of a protein that you had wanted to make in the first place.
What enzyme cuts and seals DNA?
HaeIIIrestriction enzymes
A molecule that cuts DNA molecules at a specific sequence of nucleotides?
A restriction enzyme
What is a molecule that cuts DNA molecules at a specific sequece of nucleotides?
A restriction enzyme.
What determines how DNA will be cut by a restriction enzyme?
Restriction enzymes cut DNA at specific sites called restriction sites. These restriction sites are typically 6 - 8 nucleotides in length and have a defined set of nucleotide bases. For example, the restriction enzyme Eco R1 cuts at the site: AGGTTC. Therefore, if the target DNA contains the above sequence, Eco R1 is able to cut it within the restriction site. Hence, by looking into the target site and which restriction enzymes are being used, on can make an accurate estimate of where the target DNA will be cut
What type of the enzyme that cuts the bonds of DNA molecule at the origin of replication?
restriction endonucleases
What is the term that describes sequence of DNA that a restriction enzyme finds and cuts?
Template Sequence
