What is the purpose of this experiment? Transform bacteria with the recombinant PARA-R plasmid that contains the gene of interest, red fluorescentprotein (rfp). Once transformed, the rfp gene can be expressed using the bacteria's own machinery. The PARA-R plasmid also contains a gene for ampicillin resistance (ampR), which allows us to identify transformedbacteria by their ability to grow in the presence of ampicillin.The PARA-R plasmid also contains an origin of replication (ori) to ensure the plasmid is maintained in thebacterial population, as well as the PBAD promoter and araC gene, which regulate expression of the rfp gene.How do bacteria take up DNA during transformation?Some bacteria are naturally competent, meaning they are able to take up DNA from their environment.However, not all bacteria are naturally competent, and naturally occurring transformation can be inefficient.In the lab, cells are chemically treated, and techniques like heat shock or electroporation are used to enhancethe uptake of plasmids by a variety of bacteria.Cell membranes and DNA are both negatively charged and, therefore, normally repel each other. Addingcalcium chloride neutralizes these charges and helps render the cells competent. Heat shock refers toincubating bacteria at elevated temperatures, typically 42°C, which creates holes in the membrane throughwhich DNA can enter the cell. Electroporation uses electric current to accomplish a similar result.In this simulation, the heat shock protocol will be used.DNA plasmidsare taken upBacteria expressplasmid genes andreplicateBacterialDNA->heatTransformed bacteriaare antibiotic resistantshock

Plasmids are vectors or carriers of a gene of interest that is inserted in a bacterial cell. Not all…

Answered: What is the purpose of this experiment?

Science

Biochemistry

What is the purpose of this experiment?

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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Similar questions

Find a plasmid map for pET11a and create a basic procedure for cloning a gene into this vector. Which selection method and substance would you use for this plasmid after transformation?
In gene mapping using generalized transduction, bacterial genes that are cotransduced are a. far apart on the bacterial chromosome. b. on different bacterial chromosomes. c. close together on the bacterial chromosome. d. on a plasmid.
In the formation of recombinant DNA, a restriction endonuclease cuts a bacterial plasmid to give sticky ends. The DNA segments that are to be added to the plasmid are cleaved with the same restriction endonuclease. What aresticky ends and why is it important that the target DNA and the plasmid it will be incorporated into have complementary sticky ends?
Based on the picture, why plasmids are used as a vector for DNA Recombination? What other vectors can be used?
You are attempting to clone a 3 kb gene from the bacterial sp Microbacterium foliarum into the EcoRI site of the 6.0 kb plasmid shown. If you restrict the plasmid with EcoR1, how many bands will you obtain on agarose gel electrophoresis? Compare it with a plasmid in which no gene has been inserted. Draw a representative agarose gel showing the bands, and the direction of current flow as well point out the positive and negative electrodes
Consider the ends of the DNA fragments shown below. They have been produced by digestion of a single sequence of DNA using a number of restriction endonucleases. 1. 5'A 3' 3'TTCGA5' 2. 5'G 3' 3'CAGCT5' 3. 5'AATTC3' 3' G5 4. 5'TCGAC3' 3' G5' 5. 5'GGG 3' 3'CCC 5' Which of these ends are capable of annealing and being joined by DNA ligase?
When using a conventional plasmid cloning vector containing a b-galactosidase gene, it is possible to perform a "blue-white screen" to determine which bacteria have taken up a plasmid into which a DNA fragment as been inserted, as opposed to those that have taken up just reclosed plasmid vector, by growing the transformed cells on nutrient agar plates containing the artificial b-gal substrate X-gal. Will bacteria that have taken up a plasmid into which a DNA fragment has been inserted form a blue colony or a white colony when grown on this medium? Briefly explain why these bacteria would form a colony of the color you chose.
When a 5-kb circular plasmid is digested with a restriction enzyme that has three recognition sites on the plasmid, how many bands can be visualized on an agarose gel? a)1 b)2 c)5 d)3
A 2.0kb bacterial plasmid ‘BS1030’ is digested with the restriction endonuclease Sau3A; the plasmid map is depicted in the diagram below and the Sau3A (S) restriction sites are indicated. Which of the following DNA fragments do you expect to see on an agarose gel when you run Sau3A-digested plasmid ‘BS1030’ DNA? a. 250 bp, 450 bp, 550 bp, 1.1 kb, 1.5 kb and 2.0 kb b. 2.0kb c. 250 bp, 400 bp, 450 bp, 500 bp and 550 bp d. 100 bp, 200 bp, 250 bp, 400 bp, 500 bp and 550 bp
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
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?
Explain how to identify mutant genes molecularly bytransformation with recombinant plasmids.