Download Review Topics for Final Part 2

Review topics for MCB102s 2009 Midterm #4 Part 2
***Note: I put this together before Wednesday’s lecture, but took a guess at what will be covered based on the
slides in the handout. If there are any changes or additions I will post an updated version Wednesday afternoon.***
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Prokaryotic Gene Regulation
 Negative and Positive Regulation:
— What is the difference between negative and positive regulation?
— Know the terms operon, operator, repressor, and activator
— How do allosteric effectors modulate positive and negative regulation?
— What is the difference between a cis-acting element and a trans-acting factor?
 The Lac operon:
— What genes are regulated by the Lac operon, and how do they control lactose metabolism?
— Identify the components of the Lac operon that act as a repressor, allosteric effector(s),
activator, and operator
— What are the overall effects of low or high lactose? Low or high glucose?
— What is IPTG, and what component of the Lac operon system does it mimic?
 Consensus sequences:
— What is a consensus sequence? How do variations from the consensus sequence alter the
effectiveness of a promoter?
— What is a heat shock promoter? What recognizes it?
 Attenuators (the Trp operon):
— What feature of prokaryotic transcription and translation allow for attenuation to occur?
— Understand the purpose of regions 1, 2, 3, and 4 in the Trp gene
— How does high [Trp] lead to transcriptional termination? How does low [Trp] allow for full
transcription and translation?
 SOS response:
— What types of genes does LexA repress? When would you want to turn on those genes?
— RecA protein promotes self-cleavage of the LexA repressor, but only when RecA is bound to
what?
 Regulation of ribosomal proteins and rRNA:
— When ribosomal proteins are in excess, how do they prevent synthesis of more ribosomal
proteins? Is this transcriptional or translational control?
— When amino acid levels are low, how does the stringent factor contribute to inhibition of
rRNA transcription?
 Phase variation
— Which gene of the flagellar protein regulation system is transcribed constitutively (at very low
levels)? What does the protein made from this gene do?
— How does the action of this protein control which flagellar protein is made?
Eukaryotic Gene Regulation
 Key concept: The default state of prokaryotic transcription is “on,” while the default state of
eukaryotic transcription is “off.” Why?
 Successful binding of active PolII requires the following proteins: (know these terms!)
— Transcription activator (a.k.a. transcription factor, a.k.a DNA binding transactivator)
 Bind to enhancer regions. Where can these regions be located?
— Chromatin modification proteins
 Histone Acetyl Transferases / Histone deacetylases
 SWI/SNF
 HMG (High mobility group) proteins
— Coactivators
 act as intermediaries between transcription activators and PolII
 “mediator” is the principal eukaryotic coactivator
— Basal (general) transcription factors
 We talked about these last week
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Gal4 system: which component is a transcription activator? Repressor? Enhancer? Allosteric
effector?
DNA Technology
 PCR (Pretty Cool Reaction?!?!)
— What is the purpose of each cycle of heating and cooling? Why do you need a thermostable
polymerase?
— Know how to design primers to amplify a given target region. Which strands do they bind to,
where, and in what direction?
— How can primers be used to add extra DNA to your product? How can they be used to
introduce a point mutation into a gene?
 Endonucleases
— What is the difference between blunt and sticky ends? Why are sticky ends useful?
— Why would you add a synthetic polylinker (a.k.a. multiple cloning site) to a plasmid?
— How can you use restriction enzymes to mutate a gene? (Fig 9-11)
 Bacteriophages
— What key feature of bacteriophage packaging helps to select for recombinant DNAs
containing your DNA of interest?
 BACs and YACs
— What are they? How do you put your DNA into one? Why would you use one of these
instead of a regular plasmid?
 Expression vectors: what features do these plasmids have, and why?
 Selection of Positive Clones
— How can you use antibiotic resistance genes to select for plasmids that contain your DNA of
interest? (Two different ways)
— How can you use the lacZ gene to select for positive clones? (Blue-white selection)
— Using radiolabeled probes to detect colonies of interest: you’ve got to be kidding me, no one
in their right mind would ever do this anymore when you can just sequence the DNA for $5.
Did my grandma write this book? Sigh. You should probably understand the principle
though.
 cDNA
— What is cDNA? How do you make it? What enzyme do you need? How is a cDNA library
different from a full genomic library?
 What is a fusion protein? How can you make one? Why would you want to make one? (PS, these are
incredibly useful)
 What features of our genome allow those guys on CSI to catch the right criminals?
 Microarrays
— What is printed on the chip?
— How do you make fluorescent cDNAs? What is the purpose of using more than one color?
— What can you learn from a microarray?