exam3 - sue - III exam ' GENETICS (BIO 325) (M, W, F —....

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Unformatted text preview: III exam ' GENETICS (BIO 325) (M, W, F —. 11-12 noon) Spring 2001 UNIQUE NUMBERS 46995-47015 #06101 - Time: 50 minutes Total points: [to ' Ht 1. For each of the terms in the left column, choose the best matching phrase igflie right column. (10 points) b- 1. charged tRNA 3. 2. initiation codon '\ 1.737. nonsense codon n. 4. reading frame 3. S. wobble \m 6. degeneracy of the genetic code 6 anticodon 3’. £74? g. as. aminoacyl tRNA (ti. synthetases t. 9. cap ‘9‘ 10. attenuation 1 13 fl ' ' ‘ - J1 '.-' ' 27ft . -‘ the phasein which successive triplets of nucleotides are read in mRNA '7 a tRNA to which an amino acid is attached modified guanosine nucleotide-found at the 5’ end of eucaryotic mRNAs . enzymes that attach a specific amino acid to a specific tRN A a sequence of three nucleotides in the tRNA that base pairs with a codon in the mRNA I - gene regulation involving prematnre termination of transcription AUG in a particular context most amino acids are specified by more than a single codon ' a sequence of three nucleotides in the mRNA that specifies chain termination during protein synthesis when a single tRNA' base pairs with more than one codon \3/ gr You have five T4 r11 mutants that will not grow on E. coli K (7L). You mixed two mutants together in various combinations (indicated below) and added them to E. '” ' coh’ K (a). The ability of the mixture to grow and makeplaques on E. colt“ K (at) f} - , ' was scored (indicated as a + in the table). (10 pOints) I I r .D. U" 1 ' '2 3 1 - + + 4 + 2 - - + - 3 - + - 4 - + 5 (a) What event (recombinatiou or complementation) is being scored in this analysis? ' .,_. a “Jr-w CUM‘HL (b) How many complementation groups are identified in this analysis? \p '1' ‘1. Z I Irv-J 5 11x 3 f (‘1' (c) Which mutants belong to the same complementation groups? \AFU‘ _‘9Ll003 N Oak grow“? [11 '5’4' 6' lot—lax; 'To WM A plasmid carrying genes for kanamycin-resistance (Kan’) and tetracycline- resistance (Tet') is treated with the restriction enzyme BgIII, which cleaves within the Kan' gene. The plasmid DNA is ligated with a Bglll digest of yeast DNA and the ligated mixture is used to transform E. coil. ' (10 points) (a) What antibiotic (kanamycin or tetracycline) would you use in the growth medium to ensure that each colony has the plasmid? +L“'r“‘3tl-\~L (b) How will you identify colonies in which the plasmid contains the yeast DNA qua "Md‘i-d w d0 56!. “me-3““ Lolowits in...) it» LA WVWWM+ “' / lCaMMJt-M. lfl MM. plLSMid‘. krr flu-MN: I-o hunfifi‘“: “MI 3)“— opn rnsb’t. ll. “"3 Ml, ff'n'fi-l—W’ %‘.4 I 4“ nanny} 2 Match the terms in column I with their-deft column 2: nitionfapplicationfexample given in (i) Cosmide \C, a/ , short single-stranded region at the ends of many restriction enzyme generated fragments (ii) Vectod \‘z-(wi «Ill/:— DNA copied from RNA by reverse ‘ transcriptase /C./ a collection of DNA fragments ofa given species, inserted into a vector /,. 9 / (v) cDNA [q is ,c/ a l—plasmid hybrid vector for clening large DNA fragments .a a well-defined'DNA molecule, Such as a plasmid or phage chromosome, used for cloning DNA fragments (vi) Oligonucleotide /§./; a method in which after electrophOretic separation, DNA fragments are transferre to a membrane filter for hybridization with a labeled nucleic acid probe d from a gel (vii) Probe I I 1 ends ofa DNA fragment in which all if terminal bases are paired a short, sin (viii) Western blot J DNA gle-stranded segment of used to identify specific DNA or RNA sequences by hybridization (x) Blunt enfizls C _}/ ' analysis ofproteins separated by n electrdphoresis and transferred to a membrane filter for binding with an antibody to detect a specific protein ‘/4, The following DNA sequence oon ‘ns a six base pair palindromic sequence that is recognized by a restriction endonuclease. The restriction enzyme Cleaves the DNA betWeen- the first and second nucleotide of the recognition seqUence. (10 points) _ ATC 'CTGACC -3‘ CTA ACTGG -5’ 5 ‘ - GATTTGCTAA 3 " - CTAAACGATT (a) identify the six base pair palindromic sequence that is recognized by the ‘al' restrictionenzyme. S,_f\cna-rLT' I 1" ~‘lLTF‘ h-q Cb) Cleave the DNA at this site and show the fragments obtained with their corresponding 5' and 3’ ends. s'— Pw'V 3' - T LTAtn - s‘ q’- {AATCT'I’ 3}_ [AI—q! verhangs/ sticky ends with 3’ (c) What are the ends (sticky ends with 5’ o f cleavage? overhangs! blunt ends) generated by this enzyme at the site 0 . “"“V‘3 “*5 w/ s ’ operon of E. co list are the genotypes of 5 sidase gene will ~/5. Most of'what is known about the Inc of various mutants. In the following For each strain, indicate whether the B-galacto se and whether it will be express be expressed presence of lacto (10 points) Strain I Lactose No lactose (a) I‘P*O"Z+ - __ (b) l+P*O°Zf- 4r + (c) I'lE’+O’“Zf 4, + (d) l‘i‘*O°Z+ p s (e) l"P'O+Z+ .. » When expressing eukaryotic gene pr (10 points) (a) to use cDNA instead of genomic DNA? \mkl, MJrfDfl'h 'yanwt 9M}? 7. _ k. If: w+ a“ u"- gu ergo P" (b) to fuse the cDNA with a bacterial promoter n n" Submit-L! # 14w kH-Hrm‘ rcpmdut Mm wn' ii occur abut H: W I; strains of E. cofi. in the ed in the absence of lactose. oducts in bacterial cells, why is it necessary CHI-Can. rvl- - n0 t \. .a 7. Fill in the blanks with the correct word or phrase selected from the given list. Use g each word or phrase only once (10 points) [6 Regulation of the lac Operon in E. coli is an example of Mgg'V‘t mm (b) Many bacterial promoters contain a region known as an ear W , to which repressor protein bii’ids. ' (c) Genes that are being (mama.le expressed are being transcribed all the time. (d) In the presence of glucose in the medium, there is a decreased concentration of "a. AIM? in cells of E. coli. (e) Lactose is an MAW-2r of transcription because it can bind to the repressor and prevent the repressor fiom binding to the DNA. inducer; operator; repressor; promoter; dual control; negative control only; positive control; negatively; positively; allosteric; constitutively; CAMP; lactose; r/~\ actlvator When purified E. coli RNA polymerase holoenzyme (core enzyme plus sigma a...» subunit) and ribonucleotide triphosphates are added to a-prornoter-coritainin g (I bacterial DNA in virro, transcription often initiates in a sequence~specific manner at the same site within the promoter as that initiated from the same promoter in . viva. Purified eucaryotic RNA polymerase [L however, is generally not capable of such sequence-specific initiation in vim). What factors in addition to RNA polymerase II are needed for sequence-specific initiation of eucaryotic genes? (10 points) la a (is .' HM «lo 9%)“ E’o'njyum Ill , Wt. ltrt $.qu fWH:“” (5' n * I 5, "5 C -- t , fi‘igol {lurk I'C .g ‘30“ wwLu. and M .fi‘ivmu — sat-it's :niHWO" "L "W2th 3V“. A _ _ 1:“ ,1 a i . .‘A tub-(Kalli. XML"! rim-LL“ '1 Aifih'tM mm m Paw-Hr “Ennis M cu +5. I} 9. Use the genetic code to complete the following table Assume that the reading :5 from left to right and that the columns represent transcriptional and translational aiignments ,Jf DNA double hellx QQA‘EeaTGACL‘t: u '_ u - Q; C A C_, U [2 Bid Q .__i mRNA transcribed . , it " L z 1L2. g [J G C A tRNA anticodon ~ i ' - ‘ . Trp 1: fl--_jtr_ amino acids J 1ncorporated a thermophilic bactenurn (the Taq ' J rather than a DNA Tifi gmr‘e PCR 1' some" or CL h 1qu iCi‘ilptrcLTLo-'r-/ you tic-mot l' itatCt't’sfldm-fi rotted H‘d l')i\c.Pv\[-l>_a-(ij.iu X it??? C” 17 LUC P we no Ci'tl‘lw H <3? “C1 "l ‘ W" “i Wilder I (durum/“e 1i] Which ONE ofthe f0 isolate genes?I f Ilowing is a limitation on the use ofPCR to detect and I _ known. ‘ (b) It cannot be used to amplify cDNAs. ofthe desired sequence. ,iar bacterial plasmid (pBIOBZS) has a single Hind III restriction-enzyme n the middle of a tetracycline-resistance gene (tetP‘). Yeast genomic DNA is Jested with Hind ill, and a library is made in pBIOBZS. Probing reveals that alone 15 contains a specific yeast gene of interest. Clone 15 is studied by reStriction analysis with Hind 1H and another restrictiOn enzyme, EcoRV. The ethidium bromide-stained electrophoretic gel shows bands as illustrated in the diagram below [the control was plasmid 13810325 with no insert). The sizes of the restriction fragments (in kilobaSes) are shown alongside. \\ \ Ni? \\§ .- F-r‘indlil EcoRV Hmle and EcoRV C, Control 13 Control 13 Control 1‘3 w-y/ [—l__l'_l |"_L__l_i [‘7 F"l \. i are? earn-1; " Draw a restriction map of the plasrnid pBIOBZS with no insert and ofclone lS (pBlOS25 with the insert), showing the sites for the restriction enzymes Hind Ill and EeoRV and the approximate positio%of the tetR gene. B .J'\ _ f 1 4 3‘5 699% »- xv- i brrfifrgg ____ __.i ' if) L‘)l(:‘ .. r" _/' l3. {it} The/restriction enzyme P5! 1 recognizes the following sequence in DNA and cleaves the DNA between-nucleotide Sand 6 ofthe recognition sequence. Show the ends generated by Psi l digestion and mention whether they (sticky) or blunt (flushed) are staggered lithe}: are staggered, what is the overhang (5‘ or 3‘)? ...
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exam3 - sue - III exam ' GENETICS (BIO 325) (M, W, F —....

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