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LAC OPERON (INDUCIBLE SYSTEM)
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1. Are the following E. coli cells constitutive or inducible for the z gene a. i o z b. i o z c. i oc z d. i oc z e. is o z f . iQ o z 2. Determine whether the following lac operon merozygotes are inducible or constitutive for the z gene. a. i o z / F i o z b. i o z / F i o z c. i o z / F i o z d. i oc z / F i o z e. i o z / F i oc z 3. You have isolated a repressor for an inducible operon and have determined that it has two different binding sites, one for the inducer and one for the operator. Mutants of the repressor result in three different phenotypes as far as binding is concerned. What are these phenotypes 4. An E. coli strain is isolated that produces galactosidase (lac z) and permease (lac y) constitutively. Provide two possible mutations that could cause this phenotype, and then describe how each mutation would behave in a partial diploid in which the second operon is wild-type for the entire lac system. 5. You have isolated two E. coli mutants that synthesize -galactosidase constitutively.
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* Answers to selected exercises and problems are on page A-16.
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a. If these mutants affect different functions, in what two functions could they be defective b. You can make a partial diploid of the mutants with the wild-type. What result do you expect for each mutant 6. A hypothetical operon has a sequence of sites, Q R S T U, in the promoter region, but the exact location of the operator and promoter consensus sequences have not been identi ed. Various deletions of this operator region are isolated and mapped. Their locations appear as follows, with a / representing a deleted region.
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Q Deletion 1 2 3 4 5 //////// //////// //////// ///////////////// /////// R S T U
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Deletions 3 and 4 are found to produce constitutive levels of RNA of the operon, and deletion 1 is found to never make RNA. Where are the operator and promoter consensus sequences probably located
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7. Describe the role of cyclic AMP in transcriptional control in E. coli. 8. Operon systems exert negative control by acting through inhibition. The CAP system exerts positive control because it acts through enhancement of transcription. Describe how an operon could work if it were dependent only upon positive control.
Tamarin: Principles of Genetics, Seventh Edition
III. Molecular Genetics
14. Gene Expression: Control in Prokaryotes and Phages
The McGraw Hill Companies, 2001
Exercises and Problems
9. J. Beckwith isolated point mutations that were simultaneously uninducible for the lac, ara, mal, and gal operons, even in the absence of glucose. Provide two different functions that could be missing in these mutants.
TRP OPERON (REPRESSIBLE SYSTEM)
LYTIC AND LYSOGENIC CYCLES IN PHAGE
10. Construct a merozygote of the trp operon in E. coli with two forms of the rst gene (e gene: e1, e2) in the operon. Describe the types of cis and trans effects that are possible, given mutants of any component of the operon. Can this repressible system work for any type of operon other than those that control amino acid synthesis 11. The tryptophan operon is under negative control; it is on (transcribing) in the presence of low levels of tryptophan and off in the presence of excess tryptophan. The symbols a, b, and c represent the gene for tryptophan synthetase, the operator region, and the repressor but not necessarily in that order. From the following data, in which superscripts denote wild-type or defective, determine which letter is the gene, the repressor, and the operator ( is tryptophan synthetase activity; is no activity).
Strain 1 2 3 4 5 6 7 Genotype a a a a a a a b b b b b b b c c c c c c c Tryptophan Absent Tryptophan Present
14. What is the fate of a phage entering an E. coli cell that contains quantities of repressor What is the fate of the same phage entering an E. coli cell that contains quantities of the cro-gene product 15. Describe the fate of phages during the infection process with mutants in the following genes: cI, cII, cIII, N, cro, att, Q. 16. What is the fate of phages during the infection process with mutants in the following areas: oR1, oR3 , pL , pRE , pRM , pR , tL1, tR1, nutL, nutR 17. What are the three different physical forms that the phage chromosome can take 18. How does ultraviolet light (UV) damage induce the lytic life cycle in phage 19. The prophage is sometimes induced into the lytic life cycle when an Hfr lysogen (lysogenic cell) conjugates with a nonlysogenic F cell. How might induction come about in this instance 20. A temperature-sensitive mutant of the cI gene has been isolated. At 30 C the cI repressor binds DNA, but it cannot bind DNA at 42 C (it denatures). What is the consequence of incubating E. coli that are lysogenic for this mutant at 42 C 21. The mutant in problem 20 is heated to 42 C for ve minutes, cooled to 30 C, and grown for one hour so that the cells divide several times.The temperature is then raised to 42 C, and you wait for lysis. Many of the cells are not lysed and are in fact able to form colonies. Explain these results.
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