Applications of PCR Consider this restriction enzyme (RE) named Hindlll 5...A AGCTT... 31 3... TTCGAA...5' 1. Provide a hypothetical double-stranded DNA sequence and scan the existence of these cut sites. Use different combinations of the RE in your chosen sequence to provide the fragment sizes. You may choose to give linear or circular DNA to the sequence. When the target sequence is in circular or linear DNA, explain how the fragment sizes can differ.

Applications of PCR Consider this restriction enzyme (RE) named Hindlll 5...A AGCTT... 31 3... TTCGAA...5' 1. Provide a hypothetical double-stranded DNA sequence and scan the existence of these cut sites. Use different combinations of the RE in your chosen sequence to provide the fragment sizes. You may choose to give linear or circular DNA to the sequence. When the target sequence is in circular or linear DNA, explain how the fragment sizes can differ.

Biology: The Dynamic Science (MindTap Course List)

4th Edition

ISBN:9781305389892

Author:Peter J. Russell, Paul E. Hertz, Beverly McMillan

Publisher:Peter J. Russell, Paul E. Hertz, Beverly McMillan

Chapter18: Dna Technologies: Making And Using Genetically Altered Organisms, And Other Applications

Section18.1: Key Dna Technologies For Making Genetically Altered Organisms

Problem 2SB

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In a typical microbiology laboratory, reasons for no bands from a gel of a polymerase chain reaction may bedue to errors relating to omission of ingredients in the reaction mix and absence of the target sequence inthe template DNA. Based on (i) primer problem and (ii) purity/potential contamination of the target sequence, explain the reason for non-appereance on bands.
Complete the following problems. Restriction enzymes (REs), which cut D NA at specific sequences, are classic tools in molecular biology. Because of their specificity in cutting DNA, REs can be used to "map" DNA sequences by analyzing the fragments generated upon restriction digest, as in the example shown in Figure 1. Your task is to study the circular plasmid, pMBBS, through restriction digests. You subjected the pMBBS plasmid to complete digestion by different combinations of three REs (EcoRI, BamHI, and Xhol), and analyzed the results on an agarose gel, shown below. Using the data you can glean from this gel, answer the questions that follow. EcoRI BamHI Xhol *The same total amount of DNA was loaded in each lane. 1. What is the total size of the pMBBS plasmid in bp? Answer: bp + DNA size ladder 2. How many cut sites on the PMBBS plasmid does each RE have? EcoRI: BamHI: Xhol: 3000 bp 2500 bp -2000 bp -1500 bp 1200 bp 1000 bp 900 bp 800 bp 700 bp 600 bp 500 bp 400 bp 300 bp 200 bp…
Preparing plasmid DNA (double stranded, circular) for Sanger sequencing involves annealing a complementary, single-stranded oligonucleotide DNA primer to one strand of the plasmid template. This is routinely accomplished by heating the plasmid DNA and primer to 90°C and then slowly bringing the temperature down to 25°C. Why does this protocol work? What enzyme is used and what other components are required in the sequencing reaction? How does the Sanger method determine the sequence?
ull rain LTE 11:53 © 1 0 A 10% Done #1 Mol Bio Restriction Analysi... Complete the following problems. Restriction enzymes (REs), which cut D NA at specific sequences, are classic tools in molecular biology. Because of their specificity in cutting DNA, REs can be used to "map" DNA sequences by analyzing the fragments generated upon restriction digest, as in the example shown in Figure 1. Your task is to study the circular plasmid, pMBBS, through restriction digests. You subjected the PMBBS plasmid to complete digestion by different combinations of three REs (EcoRI, BamHI, and Xhol), and analyzed the results on an agarose gel, shown below. Using the data you can glean from this gel, answer the questions that follow. EcoRI BamHI Xhol DNA size ladder 600 bp 500 bp 400 bp 300 bp 200 bp 100 bp VC 100bp Plus DNA ladder from Vivantis Technologies *The same total amount of DNA was loaded in each lane. 1. What is the total size of the PMBBS plasmid in bp? Answer: bp 2. How many cut sites on the…
Explain TWO (2) differences between two commonly used ligases; F. coli DNA ligase and T4 DNA ligase.
Given the following double-stranded fragment of DNA: 5'- ACTTGGCAGGCCTTCGATCC-3' 3'- TGAАССGTCСGGAAGCTAGG-5' A hypothetical restriction endonuclease recognizes a 6bp sequence with two-fold symmetry (typical for restriction enzymes) found in this fragment and catalyzes cleavage of this DNA on both strands between GG nucleotides within the recognition sequence. This nuclease exhibits b-type cleavage (atypical for restriction enzymes). Draw the double-stranded sequence of each fragment after cleavage showing any phosphates left on the ends.
Consulting Table 4.1, determine the length and the nature (59 or 39) of the overhang (if any) created by the following restriction endonucleases: a. Alul b. Bglll с. Clal d. Кpnl e. Mbol f. Pyul g. Not! Table 4.1 Recognition Sequences and Cutting Sites of Selected Restriction Endonucleases Enzyme Recognition Sequence* Alul AGICT BamHI GIGATCC Bgl| AIGATCT Clal ATICGAT EcoRI GIAATTC Haelll GGICC GTPY I Pu A C ALAGCIT Hindll Hindll Нраll CICGG Kpnl GGTACIC Mbol ĮGATC Pstl CTGCALG Pvul CGATICG Sall GITCGAC Smal CCCIGGG Xmal CICCGGG Notl GCIGGCCGC *Only one DNA strand, written 5'3' left to right is presented, but restriction endonucleases actually cut double-stranded DNA as illustrated in the text for EcoRI. The cutting site for each enzyme is represented by an arrow.
Consider a partial restriction digestion, in which genomic DNA is exposed to a small, limiting amount ofa restriction enzyme for a very short period of time.a. Would the resultant fragments be longer or shorteror the same size as those produced by a completedigestion?
1. Briefly explain why the total size of the pMBBS plasmid in the Restriction Enzymes practical is 3000bp (base pairs) 2. Briefly explain why the cut sites on the pMBBS plasmid in each Restriction Enzymes (EcoRI, BamHI, and XhoI) just 1 each 3. Briefly explain what led to the 5 fragments formed which linked to the 500bp, 1000bp, 1500bp, 2000bp, 2500bp sizes
1)Explain why there is a part of the Lacz gene in pUC18119 and also how the natural biochemical process wherein it normally functions is used during cloning by molecular biology. 2)Write down the basepairs of double-stranded ONA that is generated at the joint between a plasmid that is digested with Xhol and a DNA fragment that is digested with Sall and joined by DNA-ligase. Indicate the 5'- and 3'- termini of each strand.
. Propose a mechanism by which a type II topoisomerase could use the energy of ATP hydrolysis to scan a large DNA molecule and, thereby, to direct that the enzyme will catalyze largely “disentan- gling" reactions (decatenation and unknotting).
1. A DNA sequence 10 kb long contains one site at which it may be cut by the restriction enzyme Pstl. The same 10 kb long sequence also contains one site that can be cut by restriction enzyme Bg/II. Three separate fractions of this DNA were treated separately, as follows: 1) digestion by Pstl; 2) digestion by Bg/II; 3) digestion by both enzymes. The three treated fractions were then subjected to gel electrophoresis. The following table shows the lengths of the restriction fragments associated with each treated fraction. From the result, draw a restriction map that shows the Pst and the Bg/II cutting sites in the 10 kb DNA. Enzyme in mixture Pstl BgIII Pstl and BgIII Fragment Lengths (kb) 6.4 and 3.6 8.8 and 1.2 5.2, 3.6 and 1.2