The lac Operon Laboratory

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The lac Operon Laboratory
Purpose Learning to analyze an example of gene
expression Example lac operon Type of control
negative inducible
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Todays Laboratory

• Work on mutational analysis problem of lac
  operon page 3-24

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Lac OperonNegative Control and Induction
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Repressor has two binding sites.
Lactose binding site
DNA binding site
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The promoter needs to be in an ideal conformation
for RNA polymerase.
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An overview of regulation of the lac operon
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Let review a few terms and then look at the lab
problem (table) on page 3-24 important terms of
DNA control.
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3 important terms of DNA control.

• Constitutive activity
• Trans acting
• Cis acting

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Examples of cis and trans acting elements
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Cis acting Oc mutation
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Trans acting I- mutation
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Trans acting Is mutation
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Laboratory Exercise
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Haploid and Heterozygous Diploid Mutants

• Predict if you expect to obtain
• transcription/translation of the lacZ and lacY
  genes when lactose is absent or present for the
  E.coli.

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Try out your ability to reason through the
questions.
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PROTEIN PRODUCT ?-galactosidase Permease
Lactose No lactose Lactose
IPOZY -
I-POZY
IP-OZY
IPOcZY

IPOcZY-/ IPOZ-Y
IPOcZ-Y/ I-POZY-
ISPOZY-/ I-POZ-Y
IP-OcZY/ IPOZ-Y-

No Lactose
-

IPOZ-Y-/ I-POZY
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The next slides are a key to the mutations.
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Experimental tests on the repressor

• I cells synthesize full levels of the lac
  enzymes only in the presence of an inducer
• I- mutations NO DNA BINDING ACTIVITY,
  TRANSCRIPTION OCCURS EVEN IN ABSENCE OF INDUCER!
• Is mutation NO LACTOSE BINDING ACTIVITY but has
  DNA BINDING ACTIVITY. NO TRANSCRIPTION EVEN WITH
  INDUCER!

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Experimental mutations to the repressor
I sinducer binding region mutated Inhibits
transcription, even with inducer
Lactose binding site
DNA binding site
I - DNA binding site mutated, prevents
binding, allows transcription
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The End.