BIOL 1406 Discussion Questions Biotechnology 1 What is genetic recombination 2 Why is genetic recombination important 3 In nature what are the 2 basic steps involved in producing recombinant DNA 4 Describe the natural mechanisms for producing recombinant DNA in prokaryotes 5 Describe the natural mechanisms for producing recombinant DNA in eukaryotes 6 What are restriction endonucleases also called restriction enzymes 7 What is DNA ligase 8 Explain how restriction enzymes and DNA ligase are used to construct artificially recombined DNA 9 Why are scientists interested in producing multiple copies of DNA fragments 10 Describe how molecular cloning gene cloning is used to produce multiple copies of a DNA fragment Be sure to explain the role of vectors in this process 11 List and compare at least 3 different vectors that are being used to introduce foreign genes into host cells 12 Describe how the polymerase chain reaction PCR is used to produce multiple copies of a DNA fragment Be sure to list and describe the 3 basic steps of PCR 13 What is a plasmid 14 What is a recombinant plasmid 15 How do scientists use antibiotic resistance and ability to metabolize X gal to identify bacterial clones that have absorbed a recombinant plasmid 16 How do scientists screen all clones that have absorbed a recombinant plasmid to find the one s that contains a particular gene of interest 17 Explain the difference between genomic libraries and cDNA libraries 18 How is a cDNA library produced 19 If you want to make bacterial cells that can manufacture a particular human protein why is it important to insert cDNA rather than the original genomic DNA into the bacterial cells 20 How do scientists produce DNA fragments 21 If DNA from 2 individuals is treated with the same restriction enzyme why do we get different sized fragments 22 How can we determine the size of the fragments that are produced when we treat DNA from 2 individuals with the same restriction enzyme 23 How can a DNA fragment containing a particular nucleotide sequence can be isolated and identified from a sample containing many different DNA fragments 24 What are restriction fragment length polymorphisms 25 What is a DNA fingerprint 26 Explain how RFLP analysis is being in the field of forensics 27 Explain how RFLP analysis is being for genetic screening 28 What is DNA sequencing 29 Describe the method of sequencing that was developed by Fredrick Sanger 30 Why is DNA sequencing important 31 Describe at least 4 ways genetic technology can be used for human benefit 32 What is a subunit vaccine 33 How does a subuit vaccine work to provide immunity to disease 34 Describe the procedure used to construct a subunit vaccine for herpes simplex 35 How does a DNA vaccine differ from a subunit vaccine 36 Describe the procedure for preparing a DNA vaccine and explain how a DNA vaccine works to provide immunity to disease 37 What is gene therapy 38 List a few diseases that scientists have attempted to treat with gene therapy and discuss some obstacles to successful gene therapy 39 What are transgenic organisms 40 Describe some transgenic organisms that have been created for human benefit 41 Discuss some potential problems associated with genetic technology 42 The diagram below shows a segment of DNA with a total length of 4 900 base pairs The arrows indicate reaction sites for restriction enzymes enzyme X and enzyme Y Enzyme X DNA Segment Length base pairs 400 Enzyme Y 500 Enzyme X 1 200 Enzyme X 1 300 1 500 A Explain how the principles of gel electrophoresis allow for the separation of DNA fragments B Describe the results you would expect from the electrophoretic separation of fragments from the following treatments of the DNA segment above Assume that the digestions occurred under appropriate conditions and went to completion I DNA digested with only enzyme X II DNA digested with only enzyme Y III DNA digested with enzyme X and enzyme Y combined IV Undigested DNA C Explain both of the following 1 The mechanism of action of restriction enzymes 2 The different results you would expect if a mutation occurred at the recognition site for enzyme Y 43 Construct a restriction map of a linear fragment of DNA using the following data Your map should indicate the relative positions of the restriction sites along with distances from the ends of the molecule to the restriction sites and between restriction sites The Biology Place Sizes of Fragments DNA bp uncut DNA 10 000 DNA cut with EcoRI 8000 2000 DNA cut with BamHI 5000 5000 DNA cut with EcoRI BamHI 5000 3000 2000 DNA uncut DNA DNA cut with EcoRI DNA cut with HindIII DNA cut with BamHI DNA cut with EcoRI HindIII DNA cut with EcoRI BamHI DNA cut with HindIII BamHI Sizes of Fragments bp 900 700 200 600 300 500 350 50 600 200 100 500 200 150 50 500 250 100 50 45 In a molecular biology laboratory a student obtained competent E coli cells and used a common transformation procedure to induce the uptake of plasmid DNA with a gene for resistance to the antibiotic kanamycin The results below were obtained a On which petri dish do only transformed cells grow b Which of the plates is used as a control to show that nontransformed E coli will not grow in the presence of kanamycin c If a student wants to verify that transformation has occurred which of the following procedures should she use d During the course of an E coli transformation laboratory a student forgot to mark the culture tube that received the kanamycin resistant plasmids The student proceeds with the laboratory because he thinks that he will be able to determine from his results which culture tube contained cells that may have undergone transformation Which plate would be most likely to indicate transformed cells 46 Below is a plasmid with restriction sites for BamHI and EcoRI Several restriction digests were done using these two enzymes either alone or in combination a Which lane shows a digest with BamHI only Explain your reasoning b Which lane shows a digest with EcoRI only Explain your reasoning c Which lane shows a digest with BamHI and EcoRI only Explain your reasoning 47 After undergoing electrophoresis the gel in the figure below shows the RFLP analysis of DNA samples obtained from a crime scene Blood stains on a suspect s shirt B were analyzed and compared with blood from the victim V and from the suspect S Are the blood stains on the shirt from the victim or from the suspect Explain 48 Restriction endonucleases are widely used in recombinant DNA