Microbiology: An Introduction
12th Edition
ISBN: 9780321929150
Author: Gerard J. Tortora, Berdell R. Funke, Christine L. Case
Publisher: PEARSON
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Textbook Question
Chapter 9, Problem 3R
Some commonly used restriction enzymes are listed in Table 9.1 on page 248.
- a. Indicate which enzymes produce sticky ends.
- b. Of what value are sticky ends in making recombinant DNA?
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After restriction enzymes cut, they contain unpaired bases. Type II restriction enzymes leave ends that may be 5' overhanging, 3' overhanging, or blunt. In all cases each end is left with a 3' OH and a 5' phosphate. All blunt ends, and any complementary overhanging ends may be re-ligated with T4 DNA ligase, as long as at least one 5'- phosphate is present.
In the tables below G^AATTC means that the end after cutting with enzyme will be:
-----G 3'
-----CTTAA 5'
GTGCA^C means that the end will be:
-----GTGCA 3'
-----C 5'
Which RE’s from table below have a 5’ overhang? Which ones have a 3’ Overhang?
AccI
GT^CGAC
BamHI
G^GATCC
ClaI
AT^CGAT
NsiI
ATGCA^T
PstI
CTGCA^G
BglII
A^GATCT
TaqI
T^CGA
Restriction sites are palindromic; that is, they read the same in the5' to 3' direction on each strand of DNA. What is the advantage ofhaving restriction sites organized this way?
If the following is a restriction enzyme: Sma I,
A. What is the first letter represent?
B. What do the next two letters represent?
C. What does the roman numeral represent?
Chapter 9 Solutions
Microbiology: An Introduction
Ch. 9 - Compare and contrast the following terms: a. cDNA...Ch. 9 - Differentiate the following terms. Which one is...Ch. 9 - Some commonly used restriction enzymes are listed...Ch. 9 - Suppose you want multiple copies of a gene you...Ch. 9 - Which enzyme makes the smallest fragment...Ch. 9 - Describe a recombinant DNA experiment in two or...Ch. 9 - List at least two examples of the use of rDNA in...Ch. 9 - You are attempting to insert a gene for saltwater...Ch. 9 - How does RNAi silence a gene?Ch. 9 - Prob. 10R
Ch. 9 - Restriction enzymes were first discovered with the...Ch. 9 - The DNA probe, 3-GGCTTA, will hybridize with which...Ch. 9 - Which of the following is the fourth basic step to...Ch. 9 - The following enzymes are used to make cDNA. What...Ch. 9 - If you put a gene in a virus, the next step in...Ch. 9 - You have a small gene that you want replicated by...Ch. 9 - Pieces of human DNA stored in yeast cells. a....Ch. 9 - A population of cells carrying a desired plasmid....Ch. 9 - Self-replicating DNA for transmitting a gene from...Ch. 9 - A gene that hybridizes with mRNA. a. antisense b....Ch. 9 - Design an experiment using vaccinia virus to make...Ch. 9 - Why did the use of DNA polymerase from the...Ch. 9 - The following picture shows bacterial colonies...Ch. 9 - Prob. 1CAECh. 9 - Using the restriction enzyme ECORI, the following...
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- please do only part D .arrow_forwardA plasmid DNA and a linear DNA (both of the same size) have one site for a restriction endonuclease. When cut and separated on agarose gel electrophoresis, plasmid shows one DNA band while linear DNA shows two fragments. Explain.arrow_forwardOn the gel shown below are four DNA samples. Samples A to C are taken from tissues of landslide victims that are being identified, while sample D came from a hair sample brought by a mother looking for the remains of her son. (see img) i. If similar band patterns in a gel are created using the same restriction enzyme, what does that tell you about the DNA sequence of the samples? ii. In sample C, only two fragments were created. How many restriction sites (regions where enzymes cut) are present in sample C?arrow_forward
- A molecule of double-stranded DNA that is 5 million base pairs long has a base composition that is 62% G + C. How many times, on average, are restriction sites for the following restriction enzymes likely to be present in this DNA molecule? a. HpaII (recognition sequence is CCGG)arrow_forwardA molecule of double-stranded DNA that is 5 million base pairs long has a base composition that is 62% G + C. How many times, on average, are restriction sites for the following restriction enzymes likely to be present in this DNA molecule? a. HindIII (recognition sequence is AAGCTT)arrow_forwardThe sequences below indicated the 6bp recognition site for the restriction enzyme EcoRI. The lines indicate the sites where the enzyme will cut each strand. 1). write the sequence and structure of the two DNA pieces after the enzyme cuts (hydrogen bonds holding the strands together between the lines are broken after enzyme cuts) 2). indicate whether EcoRI generates blunt or sticky overhangs 5'- G I A A T T C - 3' 3' - C T T A A l G - 5'arrow_forward
- When circular DNA is sequenced, the nucleotide base pairs are numbered starting from a fixed position on the DNA, all the way around, usually in a clockwise manner. a DNA molecule that is 3133 base pairs long is digested with RsaI restriction enzyme recognition sites at base numbers 366, 1534, and 2207. What are the sizes of the DNA fragments that will be produced after the DNA is digested with RsaI?arrow_forwardA molecule of double-stranded DNA that is 5 million base pairs long has a base composition that is 62% G + C. How many times, on average, are restriction sites for the following restriction enzymes likely to be present in this DNA molecule? a. BamHI (recognition sequence is GGATCC)arrow_forward1) Restriction enzymes come in a concentration of U/ml, and it is recommended that 1U be used for each ug of DNA to be digested. If an enzyme you want to use comes in at 22,000 U/ml, and you want to digest 5 ug of DNA. How much volume will you have to use for the reaction and how will you be able to measure it with the pipettes we have in the lab? 2) An enzyme for ligation (Cip) comes at a concentration of 16000 units/ml. How many units will there be in 10 ul?arrow_forward
- You have a recombinant plasmid containing a vector and a segment of foreign DNA, both equal sizes. Draw a picture of this recombinant plasmid labeling foreign and vector regions. Where the foreign DNA meets the vector, there is a cut site for restriction enzyme ABC1. When the recombinant plasmid is cut by ABC1, how many fragments do you expect to be produced? Identify these fragments.arrow_forwardThe figure below shows the recognition sequences and cleavage positions of three restriction enzymes.You plan to ligate DNA from two different sources. The target DNA is digested with BamHI,and the insert DNA is digested with BglII, and the resulting fragments mixed and incubatedwith DNA ligase. a) Write out the sequence (in double-stranded format) of the longest insert fragment that will result after BglII digestion, ensure the nature of the overhangs is clear.b) Write out the sequence (in double-stranded format) of the ligation product, with the insert fragment joined into the BamHI site of the target DNA. Use black for target sequences, and blue for insert sequences. c) Assume the ligation reaction was successful and you have generated a recombinant DNAmolecule. Which of the three enzymes listed above can be used to excise the insert DNAfrom the target? Motivate your answer.arrow_forwardRestriction endonucleases are bacterial enzymes that cleave duplex (double-stranded) DNA at specific nucleotide sequences. The mode of replication of the animal virus SV40 has been investigated by using restriction endonucleases that cleave SV40 DNA into a number of unique segments. Like most viruses, SV40 DNA is circular. The map positions of the 11 fragments produced by a pair of restriction endonucleases are shown on the next page. Immediately following a 5 or 10 minute pulse of radioactively labeled thymidine, labeled SV40 molecules that have completed replication during the pulse are isolated. These newly replicated DNA molecules are digested by the restriction endonucleases and the resulting fragments are analyzed for the relative amounts of pulse label they contain. The results are in the table below. Assume that at the time the label was added there was a random population of replicating SV40 DNA molecules in all possible stages of synthesis. From the information given below,…arrow_forward
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