Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

BISC 320 > Analysis of Proteins and Nucleic Acids > Flashcards

Analysis of Proteins and Nucleic Acids Flashcards

1
Q

What are antibodies composed of?

A

modular domains

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What is the structure of antibodies?

A

modular domain structure - 3 domains

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

What do antibodies contain?

A

heavy and light chains (2 each) that are each composed of alternative gene segments encoding parts of the individual domains

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

What are antibodies produced by?

A

B cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What are the functions of antibodies?

A
  1. localize proteins in a cell
  2. protein purification
  3. diagnostically
  4. inhibitors of protein activity
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

What is the function of T cells?

A

produce similar proteins to distinguish self from non-self in immune surveillance - New FDA treatment involves genetic engineering of T cells to fight cancer

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

How are proteins denatured?

A

by boiling in Sodium dodecyl sulfate (SDS) and beta-mercapto-ethanol

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What is Sodium dodecyl sulface (SDS)?

A

an ionic detergent

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

What is beta-mercapto-ethanol

A

a reducing agent

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

What happens to the structure of denatured proteins?

A

loses all 2’, 3’, and 4’ structures and disulfide bonds

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

What is the function of SDS?

A

coats proteins uniformly with negative charge, and thus the amount of SDS is proportional to the length of the protein

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

T or F: SDS-coated proteins have roughly equal mass to charge ratios

A

true

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

What is PAGE?

A

SDS-Polyacrylamide Gel Electrophoresis (PAGE)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

How do proteins migrate in PAGE?

A

negatively charged, SDS-coated proteins will migrate to the positive electrode

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What is the rate at which molecules move in PAGE?

A

at a rate according to the mass:charge

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

How do SDS and BME interact with proteins?

A

completely denature proteins and give them all a similar mass:charge

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

What will the gel matrix determine?

A

the rate of migration with the rate being inversely proportional to the mass

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

How are proteins separated?

A

by size

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

How are proteins first separated in immunoblotting?

A

separated by SDS-PAGE (1D or 2D)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

How are proteins blotted in immunoblotting?

A

transferred by electrophoresis to a membrane

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

What happens to the antibody after it recognizes the protein of interest (the primary antibody, 1)?

A

incubated with the blot, and excess unbound antibody is washed away

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

What happens to the blot after it is recognized?

A

incubated with an antibody (secondary, 2) that recognizes the constant fragment (Fc) of the 1 antibody

23
Q

What happens to the 2 antibody?

A

linked to an enzyme and produces a fluorescent of chemiluminescent reaction in the presence of the appropriate substrate

24
Q

How are acidic and basic groups charged?

A

charged at pH above (acidic) or below (basic) their pKa

25
Q

T or F: uncharged proteins will migrate in an electric field?

A

false, charged proteins

26
Q

How do proteins move in a pH gradient?

A

proteins will migrate until they have no net charge will stop; this is the isoelectric point

27
Q

What is the first step in two-dimension gel-electrophoresis?

A

proteins are separated according to their isoelectric points in a tube containing a pH gradient across which a charge is applied

28
Q

When do proteins stop migrating in gel electrophoresis?

A

no net charge and then no longer will move in the electric field

29
Q

How do charged molecules move in agarose gel electrophoresis?

A

move in an electric field toward the oppositely charged pole; in the gel apparatus a buffer conducts the current

30
Q

How are DNA and RNA charged?

A

uniformly charged, so mass ratio is constant. Hence, they will migrate at same rate in electric field

31
Q

Why do gel matrix allows separation on size?

A

due to the pore size of matrix

32
Q

What sizes alter rate of movement?

A

any topology, closed circle, supercoiled circle, secondary structures of ssDNA, or RNA)

33
Q

What analysis can be done in denaturing conditions?

A

ssDNA and RNA analysis

34
Q

What allows DNA to move in an electric field?

A

negative charge of DNA and RNA

35
Q

What are used to separate DNA molecules?

A

Agarose (for DNA > 500bp) or polyacrylamide (for DNA < 1000bp) matrices are used to separate DNA molecules

36
Q

How does linear DNA migrate?

A

uniformly based on size (distance traveled is inversely proportional to its size

37
Q

What is done before gel electrophoresis can perform on DNA?

A

to determine their size, DNA molecules are first cleave to make them linear

38
Q

How is DNA vizualized

A

with a stain

39
Q

What protects the host genome from foreign DNA?

A

restriction-modification systems

40
Q

What does bacteria produce?

A

produce a restriction endonuclease also must produce a DNA modification enzyme (a methylase) to protect their own DNA from digestion

41
Q

What does methylation in the recognition sequence prevent?

A

prevent the binding of the restriction enzyme to the sequence, thereby preventing cleavage

42
Q

How do restriction enzymes bind to DNA?

A

as dimers

43
Q

What is the function of restriction enzyme dimers?

A

recognize palindromic DNA sequences (two half sites) and cleave DNA symmetrically on both strands of the DNA

44
Q

What produces restriction enzyme?

A

produced by bacteria protect against invading DNA (e.g. phage infection)

45
Q

What does the length of recognition sequence determine?

A

the approximate cleavage frequency

46
Q

What does DNA base composition affect?

A

affect frequency of cleavage (e.g. AT- rich versus GC-rich DNA

47
Q

What is the frequency of a specific sequence in DNA?

A

f = (0.25)^n; n = length of sequence

48
Q

Is sequence composition random?

A

not random; some sequences are more or less abundant than expected on random basis

49
Q

What is the function of an electrophoretic mobility shift assay (EMSA)?

A

used to determine the interaction between protein and DNA molecules

50
Q

What is a Non-denaturing gel?

A

usually agarose, may be polyacrylamide

51
Q

What is an example of a gel-shift?

A

a DNA molecular ( the probe) moves more slowly when bound to a protein. As more protein is added, a greater proportion of DNA moves at a slower rate through the gel. Testing different protein concentrations allows the binding kinetics of the protein to be determined

52
Q

What are the four steps of Chromatin Immunoprecipitation (ChIP)

A
  1. treat live cells with chemicals (e.g. HCHO) to cross link proteins to DNA
  2. fragment DNA randomly
  3. use antibodies to immunoprecipitate specific protein along with DNA to which it binds
  4. reverse crosslinking to remove protein and purify DNA
53
Q

How are DNA identified in ChIP?

A

by PCR (ChiP), hybridization (ChIP-chip), or high-throughput sequencing (ChIP-seq)

BISC 320 (20 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions