PSwk7 - Problem
Set
Week
7
...

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Unformatted text preview: Problem
Set
Week
7
 MCB110,
Spring
‘11
 
 1.

True
or
false:
 _______
TBPs
and
TAFs
are
part
of
TFIIA

 _______
Eukaryotic
core
promoters
contain
a
‐35
and
‐10
region
 _______RNAP
II
is
responsible
for
transcribing
nearly
all
genes
 _______Eukaryotic
core
promoter
recognition
requires
3
proteins
 _______There
are
2
phosphorylation
steps
in
bacterial
transcription
initiation
 _______CpG
islands
are
commonly
found
upstream
of
housekeeping
genes
 ______
The
gel
used
in
Gel
Shift
experiments
(EMSA)
is
denaturing
 _____
DNaseI
footprinting
assay
is
an
in
vitro
way
to
test
whether
a
protein
activates
 transcription
 ______
Most
DNA
binding
domains
of
transcription
factors
are
devoid
of
arginines
 and
lysines
 ______
High
salt
washes
in
affinity
chromatography
get
rid
of
nonspecifically
or
 weakly
interacting
proteins.
 
 2.
List
the
functions
of
the
three
different
eukaryotic
RNA
polymerases.
 
 
 
 
 
 
 3.

If
you
are
given
a
candidate
RNA
Polymerase
protein
how
might
you
determine
 which
type
of
RNA
Polymerase
it
is?
 
 
 
 
 
 
 
 4.

Describe
how
Eukaryotic
RNAP
II
is
recruited
to
the
core
promoter.

How
is
this
 different
from
the
recruitment
RNAP
in
bacteria.
 
 
 
 5.

What
marks
the
1st
step
of
eukaryotic
promoter
recognition?
 
 
 
 6.

Imagine
you
are
conducting
an
in
vitro
step‐wise
assembly
of
the
RNAP
II
Pre‐ initiation
complex
at
a
core
promoter.

 A.

What
would
be
the
specific
consequence
of
leaving
TFIID
out
of
the
assembly
 reaction?
 
 
 
 B.
What
would
be
the
specific
consequence
of
leaving
TFIIE
out
of
the
assembly
 reaction?
 
 
 
 
 C.
What
would
be
the
specific
consequence
of
leaving
TFIIH
out
of
the
assembly
 reaction?
 
 
 
 

 7.

Describe
how
RNAP
II
is
regulated
during
transcription,
especially
during
 promoter
clearance
and
elongation.
 
 
 
 
 
 8.

We
have
learned
that
a
gel
shift
assay
is
one
method
that
can
be
used
to
study
a
 protein‐DNA
interaction.

What
type
of
gel
is
used
in
this
assay?

What
are
the
 advantages
and
disadvantages
to
using
this
assay?

 
 
 
 
 
 9.

When
doing
an
in
vitro
transcription
assay,
what
components
must
you
include
 in
the
reaction?
 
 
 
 
 
 10.

Imagine
you
are
doing
an
in
vitro
transcription
assay
to
look
for
the
ability
of
a
 particular
transcription
activator
to
activate
transcription
of
a
gene.

What
can
be
 used
as
a
positive
control
for
this
assay?
 
 
 
 
 11.
If
you
want
to
test
whether
a
protein
is
a
transcription
repressor,
what
 technique
we’ve
learned
so
far
in
class
would
help
you
accomplish
that?
 
 
 
 
 
 12.
How
would
you
tell
whether
a
DNA
sequence
could
be
a
bona
fide,
true
enhancer
 sequence
without
doing
any
bench
work?
 
 
 
 
 
 13.
You
discovered
a
transcription
factor
that
prefers
to
bind
promoters
that
have
a
 consensus
AT‐rich
binding
site
 a.
How
would
you
purify
this
protein?
 
 
 b.
If
there
are
three
AT‐rich
similar
but
slightly
different
binding
sites
in
the
 promoter
of
a
particular
target
gene
of
this
transcription
factor,
then
describe
two
 experiments,
one
in
vitro
and
one
in
vivo,
that
you
can
use
to
tell
exactly
which
site
 is
the
true
binding
site
 
 
 c.
What
initial
(bench)
experiment
could
you
do
to
search
for
new
direct
targets
of
 this
protein,
knowing
that
cycloheximide
is
a
commonly
used
small
molecule
that
 inhibits
translation
of
new
protein
products
(by
inhibiting
translocation
of
 ribosome)
 
 
 
 
 14.
Describe
the
three
major
DNA
binding
motifs
of
transcription
factors
that
we’ve
 covered
in
class.
What
are
some
of
the
common
features
shared
by
two,
or
among
all
 three?
 
 
 
 
 15.
How
can
you
map
(determine
the
location
of)
the
DNA
binding
module
of
a
 known
transcription
factor?
What
about
the
dimerization
module?
What
assay
 would
you
choose
and
describe
the
experimental
set
up.

Hint:
for
both
you
can
use
 an
in
vitro
assay
we
talked
about,
and
for
the
former,
you
can
also
use
an
in
vivo
 technique
that
we’ve
mentioned.
 ...
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This document was uploaded on 09/12/2011.

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