Quizzes PGM

1
Quizzes - PGM

• 2008

2
Quiz 1 24 September 2008

• 1) True or False. The first homework assignment
  is due on Friday of this week.
• 2) Research done simply for the joy of finding
  out an answer is called
• A) Basic research B) Applied research
• 3,4) Name any 2 of the three hurdles a molecular
  biologist must overcome in order study his or her
  favorite gene.
• a) have enough to work with b) identify the DNA
  that has the gene of interest c) separate the
  DNA of interest from all the other DNA.
• 5) Name either one of two types or sources of DNA
  that can be replicated to high copy number in a
  bacterial culture.
• Hint One starts with P and one starts with B
  (or P for short).
• Plasmid and bacteriophage. PCR and cloning are
  also acceptable answers.

3
Quiz 2 26 September 2008

• Remember to put last name first on the cover and
  the page on which you are writing your answer.
  Please use pen and avoid teensy weensy lettering.
• What piece of equipment do you work with if you
  are a bioinformaticist or a computational
  biologist? Computer
• What is the advantage to a bacterium of producing
  a restriction endonuclease? (1 sentence or less)
• Makes it possible for bacteria to digest DNA of
  invading pathogens so they cant replicate.
• 3) True or False Both plasmids and
  bacteriophage are easy to isolate from bacterial
  cultures.
• 4) True or False. An RE that recognizes
  5CAGCTG3 also recognizes 5GTCGAC3.
• 5) Note the following RE recognition sequence
  CACGTG.
• Write a different recognition sequence
  that is isocaudameric. Be sure to show the cut
  site.
• GACGTC or AACGTT or TACGTA
• Position of cut site and the nucleotides of the
  resulting tail must be the same. The nucleotides
  outside the cuts may change, but they must remain
  palindromic.

4
Quiz 3 29 September 2008

• Which RE would you use if you wanted longer DNA
  fragments as products of your restriction
  endonuclease digest? One that cuts at
• A) GACGTC or B) GGCGCGCC
• Where on the DNA is the negative charge located?
• On the sugar phosphate backbone, on the singly
  bonded oxygen at each phosphate. Also
  acceptable at the 5 phosphate of a DNA strand.
• True or False. During agarose gel
  electrophoresis, DNA travels toward the positive
  electrode.
• True or False. Smaller DNA molecules move
  through the gel more quickly than larger ones.
• What is different about how the charge is applied
  in OFAGE as opposed to standard agarose gel
  electrophoresis? In OFAGE, the charge oscillates
  (changes) between at least two different
  directions. In standard, the direction is
  constant.

5
Quiz 4 1 October 2008

• 
  
• 1) What is the name given to the location in a
  plasmid at which one finds a series of closely
  spaced RE sites that appear nowhere else in the
  plasmid? Multiple cloning site, polylinker.
• 2) What should be the very first step in any
  recombinant DNA project, or for that matter, any
  project? Make a plan or devise a strategy.
• 3,4) There are two major pieces of DNA that you
  need to prepare in the process of making a
  recombinant DNA construct. What are they? (Do
  not use plasmid as an answer.) Vector and
  insert.
• 5) Name the enzyme used for
• a) Nicking of RNA that is base paired with first
  strand synthesis cDNA. RNAse H.
• b) 2nd strand synthesis of cDNA. DNA polymerase
  I.
• c) Making the ends of ds cDNA blunt. T4 DNA
  polymerase.
• d) Colvalently attaching ends of DNA to each
  other. DNA ligase.

• 
  Last Name, First Name
• 1,2.
• There are two major pieces of DNA that you
  need to prepare in the process of making a
  recombinant DNA construct. What are they?
• 3,4.
• Name the enzyme used for
• A) First strand synthesis of cDNA
• B) Nicking of RNA that is base paired with first
  strand synthesis cDNA.
• C) 2nd strand synthesis of cDNA.
• D) Making the ends of ds cDNA blunt.
• 5. True or False The restriction enzyme
  recognition sequences that appear in a useful
  multiple cloning site do not appear elsewhere in
  the plasmid.

6
Quiz 5 3 October 2008

• True or False. Sticky ends can be ligated even
  when no 5 phosphate is present.
• 2. What is the adjective or name given to
  bacterial cells that have been prepared for use
  in a transformation reaction? Competent
• 3. You are attempting to clone an insert into a
  vector which contains a single drug resistance
  gene. The gene codes for ampicillin resistance.
  Following transformation, you spread the bacteria
  on a plate containing ampicillin and allow the
  bacteria to grow overnight.
• True or False This process tells you which
  bacteria contain constructs with insert.
• 4. What is the name of the enzyme that breaks
  down a lactose analog resulting in a blue color?
  Beta-galactosidase LacZ is not absolutely
  correct because its only part of the enzyme but
  was accepted as an answer.
• 5. If your strategy is to place a cDNA insert
  into the Bam HI site of a vector, what adaptors
  do you use to give your cDNA sticky ends?
• a) Pst I b) Bam HI c) you dont need
  adaptors

7
Quiz 6 6 October 2008

• If you are eligible to vote, and have not
  registered, or need to change your registration,
  please do!
• The deadline is October 20.
• Forms for registering in CA are available at
  http//www.sos.ca.gov/elections/elections_vr.htm
• Check also http//www.eac.gov/voter/Register20to
  20Vote

8
Quiz 7 8 October 2008

• In a blot hybridization protocol, what is the
  word for the labeled DNA that includes your
  sequence of interest? Probe
• Give an example of what the label might be for
  1. Radioactive 32P, 33P, 35S
• Non-radioactive biotin, fluorescein,
  digoxigenin, alkaline phosphatase, horseradish
  peroxidase
• 3) One general way to label is by chemical
  cross-linking. What is the other general way?
  Enzymatically.
• 4) What is the word that means to use heat or
  alkaline conditions to separate the two
  complementary strands of dsDNA from each other?
  Denature.
• What is the word given to blot hybridization in
  which RNA has been blotted to the membrane from a
  gel? Northern.

9
Quiz 8 10 October 2008

• In which phosphate do you want your radioactive
  label to be if you are going to label the 5 end
  of a probe?
• Alpha Beta Gamma
• 2,3) You plan to make a cRNA probe. Your student
  finds that some labeled NTP has just arrived and
  is in the freezer. You look at the package
  description closely and say, That wont work.
  Thats for someone who is kinasing. What must
  the package description have said, and why wont
  that work for cRNA labeling? Gamma labelled ATP.
• 4) DNA that functions as a primer is required
  during
• A) labeling by random priming
• B) labeling by filling in a recessed 3 end
• C) labeling by chemical cross-linking
• D) both A and B
• 5) To label the 5 end of a strand of DNA you
  will need
• A) kinase
• B) polymerase
• C) ligase

10
Quiz 9 13 October 2008

• 1. If you use a probe labeled with alkaline
  phosphatase, what kind of signal will be measured
  when you visualize your results? (There are two
  possible answers. One is more commonly correct
  than the other.) light or color
• 2. T or F When you have an X-ray done at a
  physicians laboratory, you are undergoing an
  autoradiography procedure.
• 3,4. If you label a probe with biotin, what two
  additional reagents must be added before you can
  produce a signal to visualize the locations at
  which the probe has hybridized?
• Streptavidin-conjugated enzyme such as
  horseradish peroxidase or alkaline phosphatase
• a substrate from which the enzyme produces light
  
• 5. Which allows better quantitation of hybridized
  probe, exposure to X-ray film or phosphorimaging?
  phosphorimaging

11
Quiz 10 15 October 2008

• Give one of the two major advantages of using
  bacteriophage instead of conventional plasmids
  for construction of libraries.
• More efficient introduction of DNA into cells.
• Able to accommodate a larger insert and still be
  efficient at putting DNA into cells.
• Name any two of the 3 required parts of the
  lambda phage genome that are required for the
  lytic life cycle.
• lambda left arm, lambda right arm, cos ends.
• 3. What is the name given to a clone of phage on
  an agar plate?
• Plaque
• 4. T or F. A good eukaryotic cDNA library will
  have inserts that are just as big as the inserts
  in a good genomic library.
• 5. T or F. A cDNA library made from the mRNA
  from one type of cell will be the same as a cDNA
  library made from any other type of cell.

12
Quiz 11 17 October 2008

• 1. T or F In order to remove the stuffer
  (central region) from a lambda vector, you need
  to cut the vector at the COS sites.
• 2. Which of the following digestions of the
  genomic DNA with which you plan to construct a
  library will allow you to achieve inserts that
  are predominantly the size you wish for a lambda
  replacement vector?
• a) partial digestion with a 6-hitter
• b) partial digestion with a 4-hitter
• c) complete digestion with a 6-hitter
• d) complete digestion with a 4-hitter
• 3. T or F. If one lambda right and one lambda
  left arm of a replacement vector ligate to each
  other without an insert, no plaque will form.
• 4. T or F. After ligation of inserts to arms, a
  lambda library is transformed into competent
  bacteria.
• If two lambda right (short) arms ligate to each
  other during preparation of a genomic library,
  the product will not package. Explain. Too
  short to package, and even if it did, would lack
  the part of the genome required to code for
  structural proteins in vivo. Note that the
  question was about packaging, not replication and
  construction of new phage.

13
Quiz 12 27 October 2008

• Name any one high capacity vector other than a
  cosmid. P1, PAC, BAC, YAC.
• Use one or two sentences to describe any one
  feature of a cosmid that contributes to its name.
  
• Includes lamda cos sequences so that it can be
  packaged as a phage. Includes plasmid ori so
  that it can replicate as a plasmid. Has
  antibiotic resistance gene so can be selected as
  a plasmid.
• 3) Name one prokaryotic feature of a YAC vector
  and one eukaryotic feature.
• Prokaryotic plasmid ori, bacterial drug
  resistance
• Eukaryotic centromere, telomere, yeast
  biosynthetic enzymes.
• 4) Red color, white color, and sucrose tolerance
  are different ways of indicating what? Presence
  of an insert.
• 5) What advantage do electroporation and
  packaging as phage have over transformation as a
  method of introducing DNA into bacterial cells?
  More efficient more constructs get from outside
  the bacteria to inside the bacteria.
• 6) Bonus Do any of the high capacity vectors we
  have described replicate as phage within
  bacterial cells? No.
• I see the misunderstanding with respect to P1
  and PAC, so I threw the question out.
• The term replicate as a phage implied going
  through the full phage life cycle, which doesnt
  happen with anyof the high capacity vectors that
  we have discussed.

14
Quiz 13 31 October 2008

• What is missing from ddNTPs that causes a chain
  termination whenever a ddNTP is incorporated? The
  3 hydroxyl
• How are the four bases distinguished from each
  other in current fluorescence-based sequencing
  techniques? Each base is labeled with a
  different fluorescent color.
• True or False Cycle sequencing uses the same
  DNA as template multiple times.
• True or False Sequencing requires the inclusion
  of 3 bases as dNTPs and the fourth as a ddNTP.
• In order to be sequenced correctly by
  polyacrylamide gel or capillary electrophoresis,
  DNA must be
• a) single-stranded b) denatured
  c) both

15
Quiz 14 - 3 November 2008

• Name any one way in which PCR differs from cycle
  sequencing. Possibilities PCR has 2 primers
  sequencing,only 1. PCR both strands are used as
  template sequencing, only one. PCR no ddNTPs
  are used cycle sequencing, ddNTPs are used.
  (There are other possible answers.)
• Name any one major difference between Solexa (or
  454) sequencing and conventional cycle
  sequencing. Solexa sequences millions of
  templates simultaneously conventional sequences
  one template per reaction vessel. Solexa and
  454 are good for many short reads conventional
  cycle is best for fewer longer reads. Solexa and
  454 do not depend on ddNTP to generate a series
  of fragments of different lengths. Solexa and
  454 are analyzed after each base addition
  conventional is analyzed at the end of mutliple
  extension periods.
• What is the trick used in screening a large
  library that reduces the total number of PCR
  reactions required? (One word, starts with p)
  POOL.
• True or False The more variability within the
  population in the number of STRs at a single
  locus, the more useful that locus is for
  identification purposes.
• Name a feature of the technique of PCR that makes
  it so useful for forensics.
• Template sample can be somewhat degraded. You
  dont need much template. Fairly cheap and easy.
  

16
Quiz 15 - 5 November 2008

• Name any one thing that was significant about
  yesterday in US history.
• Most of you said first African-American
  president-elect. A couple mentioned the
  record-setting voter turnout. There were other
  equally correct answers.
• If you voted in the election, about how long did
  you have to wait in line?
• 0 hours!
• Who is the president-elect of the United States?
  Senator Barack Obama.
• What right did egg-laying chickens gain in the
  state of California? The right to enough space
  to stand up, turn around, and spread their wings.
• Will there be a new puppy in the white house?
  Yes!
• PCR results can be analyzed by
• a) Gel electrophoresis b) Capillary
  electrophoresis
• c) fluorescent dye binding d) All of the
  above.
• In real time PCR, a measurement is taken of how
  much product has been made a) just once after
  all the cycles have been completed, or b) every
  time a new cycle has been completed.
• True or False Promoter sequences can be studied
  both in silico and in vitro.

17
Quiz 16 7 November 2008

• True or False ChIP determines where
  transcriptional regulatory proteins are bound in
  living cells.
• True or False ChIP tells you whether or not
  binding of a protein turns transcription on or
  off.
• What is a reporter assay meant to report?
  Efficiency of the sequence you are testing to
  drive transcription.
• If you have made a reporter plasmid and need to
  have some more, in what organism do you grow up
  additional plasmid? E. coli.
• You carry out a reporter assay. The construct
  containing -700 to -1 gives you 1000 light units.
  The construct containing -550 to -1 gives you
  3000 light units. What might you conclude about
  the region between -700 and -550? The region
  binds a repressor. Once the region is removed
  and repressor can no longer bind and be active,
  transcription increases.

18
Quiz 17 12 November 2008

• 1. True or False EMSA can be used to test the
  hypothesis that an unknown protein or proteins
  present in your cell of interest can bind to a
  specific sequence of ds DNA.
• 2. In order to perform serial deletions with
  Exonuclease III, you need
• a) 3 recessed end to prevent deletion and 3
  overhang to allow it.
• b) 3 recessed to allow Exonuclease III
  activity and 3 overhang to prevent it.
• c) It doesnt matter because both 3 recessed
  and 3 overhangs allow for Exonuclease digestion.
• 3. Name any one feature that reporter plasmids
  and expression plasmids have in common. E. coli
  origin of replication drug resistance gene for
  selection in E. coli (multiple) cloning site.
• 4,5. In a single sentence, describe the most
  important distinction between what type of
  sequence you would clone into a reporter plasmid
  vs what type you would add into an expression
  plasmid. A promoter sequence would be cloned
  into a reporter plasmid, whereas a protein coding
  sequence would be cloned into an expression
  plasmid.

19
Quiz 18 14 November 2008

• You are planning an expression system. You want
  your human protein to be expressed in E. coli.
  The protein is a protein with no equivalent in
  bacteria.
• 1-5. List any five issues that you must/may
  need to address in designing your system. (No
  paragraphs. Just a list. Complete sentences are
  OK, but not necessary.)
• Please check your Expression System lecture
  slides for multiple possible answers to the above
  question.

20
Quiz 19 17 November 2008

• True or False. It is possible to transfer an
  expression plasmid into eukaryotic cells.
• Name any 4 ways in which DNA can be transferred
  into cells. Electroporation, DNA precipitation,
  liposomes, dendrimers, biolistics,
  microinjection, viral transfer, bacterial
  transfer (plants).
• A protein that includes not only its own domains,
  but additional domains is called a fusion
  protein.
• What domain allows you to purify YFP on a
  nickel column? His tag
• Name any one way in which extra domains might be
  removed from YFP in vitro. Protease cleavage at
  a specific amino acid sequence engineered between
  domains cyanogen bromide cleavage at a unique
  methionine engineered between domains.

21
Quiz 20 21 November 2008

• True or False A targeting vector is required
  for making a knockout mouse.
• True or False Chimeric mice may be either
  germline or somatic transgenics.
• What must be included in a vector for stable
  expression in eukaryotic cells that is not
  required for transient expression? A gene for
  selection in the eukaryotic cell.
• What feature of a targeting construct gives it
  the ability to target a given genomic location?
  Regions of homology to your target gene.
• Explain how the blue mouse is similar to a
  promoter/reporter experiment. Your answer should
  have two parts.
• The blue is a reporter that reports on the
  appropriateness of expression driven by
  YFPromoter, which drives the reporter.
• ( Both the promoter and the reporter are
  transgenic, as they are in promoter/reporter
  experiments we have already discussed.)