What's the cloning process from the first step to the last? Put the steps in order based on the following options: -Cut out the gene of interest with a restriction enzyme -Transform the E. coli with the plasmid. -Ligate the backbone of the DNA from the plasmid to the gene of interest. -Allow the gene of interest to anneal at the insertion site in the plasmid. -Select colonies based on the reporting system used in the plasmid. -Cut open the multiple cloning site on a plasmid with a restriction enzyme.

What's the cloning process from the first step to the last? Put the steps in order based on the following options: -Cut out the gene of interest with a restriction enzyme -Transform the E. coli with the plasmid. -Ligate the backbone of the DNA from the plasmid to the gene of interest. -Allow the gene of interest to anneal at the insertion site in the plasmid. -Select colonies based on the reporting system used in the plasmid. -Cut open the multiple cloning site on a plasmid with a restriction enzyme.

Human Heredity: Principles and Issues (MindTap Course List)

11th Edition

ISBN:9781305251052

Author:Michael Cummings

Publisher:Michael Cummings

Chapter13: An Introduction To Genetic Technology

Section: Chapter Questions

Problem 11QP: Cloning Genes Is a Multistep Process In cloning human DNA, why is it necessary to insert the DNA...

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As you know, restriction enzymes evolved in different bacterial species independently. The adaptive significance of having a restriction enzyme is that the bacterium has the ability to cut the injected viral DNA into small segments. This destruction of viral DNA prevents the virus from taking over the bacterial cell and killing the cell. What is one benefit of using a restriction enzyme with staggered ends (such as EcoRI) to cut both the DNA insert and the plasmid? Which types of cut sites (staggered with “sticky ends” or blunt ends) are most useful in cloning DNA? Would you expect restriction enzymes in different bacteria genera (Streptococcus, Lactobacter, Escherichia) to have the same recognition sites (DNA sequences). Why or why not?
In order to construct a genome library, which statement most accurately describes the correct order of steps in the process? Fragment chromosomal DNA, digest plasmid vector, ligate genomic DNA and digested plasmid vector, transform ligation into bacteria, select for plasmid marker Fragment chromosomal DNA and plasmid vector, ligate genomic DNA and plasmid vector, transform ligation into bacteria, select for gene marker Fragment chromosomal DNA and plasmid vector, ligate genomic DNA and plasmid vector, transform ligation into bacteria, select for plasmid marker Fragment chromosomal DNA, ligate genomic DNA and plasmid vector, transform ligation into bacteria, select for plasmid marker
The following statements are true about common gene cloning procedures except: -DNA plasmids can help move around genes into other cells - Restriction enzymes are very important for cutting up and linking chunks of DNA -We make it so that genes from plants or animals are expressed in bacteria so the products can be harvested - DNA plasmids are chunks of chromosomal DNA used for cloning
What is the most logical sequence of steps for splicing foreign DNA into a plasmid and inserting the plasmid into bacterium?I) Transform bacteria with recombinant DNA molecule.II) Cut the plasmid DNA using restriction enzymes.III) Extract plasmid DNA from bacterial cells.IV) Hydrogen-bond the plasmid DNA to non-plasmid DNA fragments.V) Use ligase to seal plasmid DNA to non-plasmid DNA A. III, II, IV, V, I B. I, II, IV, III, V C. III, IV, V, I, II D. II, III, V, IV, I
Which of the following would you need to clone a human gene? Choose all that apply A restriction enzyme Human DNA A donor goat nucleus An enucleated egg A surrogate mother DNA ligase A plasmid vector (or other type of DNA vector, e.g. phage, BAC)
You’re working in a research lab, and your current task is to clone the gene that codes for tyrosinase from potatoes. You grind up some potato, extracts the DNA from it and digests the DNA with two different restriction enzymes (separately, not together): EcoRI and BamHI. You then obtain the cloning vector, pUC19, and digest it with the same two enzymes. You then run a gel which is shown to the right. You mix the potato DNA (digested with the enzyme you specified in part B) with cloning vector DNA (digested with the same enzyme). You then add the mixture to E. coli cells and carry out a transformation procedure so that the cells can each update a plasmid. You then plate the cells on a plate containing antibiotic. What antibiotic would be in the plate? Why would there be antibiotic in the plate? Be specific! Unfortunately, you don’t get a single bacterial colony to grow on the plate. Not even one! You review your procedure and realize that when mixing the digested potato DNA…
If I clone a complete eukaryotic gene, including the eukaryotic promoter region, ligate it into a plasmid, and transform it into E. coli, will I be able use the transformed E. coli to make the corresponding protein? Explain why, or why not? If you decide to do this, what would your cloning strategy be?
When making recombinant DNA, why must you use the same restriction enzyme to cut the gene of interest and the plasmid?
Cloning vector contains a multiple cloning site (MCS) or polylinker containing multiple unique restriction sites. It also contains markers that allow for rapid selection of colonies with insert-containing plasmids in them. The 1st statement is True and the 2nd statement is False. The 1st statement is False and the 2nd statement is True. Both Staments are True. Both statements are False.
In cloning human DNA, why is it necessary to insert the DNA into a vector such as a bacterial plasmid?
An important feature of restriction enzymes is that each enzyme only recognizes a specific palindrome and cuts the DNA only at that specific sequence of bases. A palindromic sequence can be repeated a number of times on a strand of DNA, and the specific restriction enzyme will cut all those palindromes, no matter what species the DNA comes from. A linear DNA molecule is represented below. The DNA is represented by one line, although in actuality, DNA has two strands. If the DNA molecule has two restriction sites, specifically two repeats of a specific palindrome sequence, A and B, for a specific restriction enzyme: How many fragments would be produced if the DNA is cut by that enzyme? Number each fragment Which fragment would be the largest? Which fragment would be the smallest?
With the use of well-illustrated diagrams, reconstruct the entire cloning process by explaining different stages of the cloning process that involves the following:a. Isolation of target DNA fragments (often referred to as inserts)b. Ligation of inserts into the plasmid, creating recombinant molecules c. Transformation of recombinant plasmids into bacteria or other suitable host for propagationd. Screening/selection of hosts containing the intended recombinant plasmid. For this stage(d), discuss the importance of a second marker that can be used for screening of genomic DNA for colonies containing the pka-1 under the principle of insertional inactivation. This should be properly explained using all the attributes of the plasmid described above.