lab techniques

Question 1

gel electrophoresis separates what

Answer 1

macromolecules (proteins, DNA, RNA)

Question 2

what is used for proteins/small molecules in gel electrophoresis

Answer 2

polyacrylamide

Question 3

what is used for larger molecules in gel electrophoresis

Answer 3

agarose

Question 4

negatively charged molecules travel toward

Answer 4

anode at bottom

in gel electrophoresis

Question 5

larger molecules move…

Answer 5

SLOWER in gel electrophoresis

Question 6

what dye is used in gel electrophoresis

Answer 6

coomassie blue stain for visualization

Question 7

native-PAGE is a method where

Answer 7

polyacrylamide gel electrophoresis method for proteins is utilized with NON-DENATURING conditions

Question 8

proteins keep their native charge and structure so that…

Answer 8

they are separated based on charge and size

in native-PAGE

Question 9

in SDS-page, a polyacrylamide gel electrophoresis method is used for

Answer 9

proteins using DENATURING conditions

Question 10

sodium dodecyl sulfate denature the…

Answer 10

proteins and give them a uniform charge

part of SDS-PAGE

Question 11

proteins separate solely on mass and then you can…

Answer 11

estimate the protein’s molecular mass

SDS-PAGE

Question 12

reducing SDS-PAGE is the same as

Answer 12

SDS-PAGE but adds a reducing agent

Question 13

reducing agent in reducing SDS-PAGE will reduce

Answer 13

disulfide bridges and result in completely denatured protein

Question 14

isoelectric focusing is where a gel electrophoresis method is used to

Answer 14

separate proteins on the basis of relative contents of acidic/basic residues

Question 15

gel has pH gradient and proteins will migrate through gel until….

Answer 15

they reach pH that matches isoelectric point

Question 16

at the pI, the protein has a neutral charge so

Answer 16

it will no longer be attracted to the anode and it will stop migrating

part of isoelectric focusing

Question 17

southern blotting

Answer 17

detection of a specific DNA sequence in a sample

Question 18

northern blotting is the detection of

Answer 18

specific RNA sequences in a sample

Question 19

western blotting is the detection of a

Answer 19

specific protein in a sample

Question 20

sanger DNA sequencing uses what method

Answer 20

chain termination method

Question 21

chain termination method uses

Answer 21

dideoxy nucleotides

Question 22

ddNTP lacks a

Answer 22

hydroxyl group on 3’ carbon on sugar ring

no more nucleotides can be added to the chain w/ the 3’ hydroxyl group missing

Question 23

chain ends with the

Answer 23

ddNTP that is marked with a particular color of dye depending on base carried

Question 24

polymerase chain reaction (PCR) is used to make

Answer 24

many copies of a specific DNA region in vitro

Question 25

key ingredients of PCR =

Answer 25

Taq polymerase primers template DNA nucleotides (DNA building blocks)

Question 26

ingredients are assembled in a tube along with cofactors needed by the enzyme and put through.....

Answer 26

repeated cycles of heating/cooling that allow DNA to be synthesized

Question 27

primer must have high...

Answer 27

GC content and either a G or C at each end

Question 28

enzyme used in PCR

Answer 28

taq polymerase

Question 29

taq polymerase is named after the

Answer 29

heat-tolerant bacterium from which it is isolated very heat-stable and most active around 70 C

Question 30

steps of PCR

Answer 30

1) denaturation (96 C) 2) annealing (55 - 65 C) 3) extension (72 C)

Question 31

PCR's cycle is repeated until

Answer 31

you have enough DNA

Question 32

chromatography separates

Answer 32

two or more molecules from a mixture

Question 33

stationary phase is

Answer 33

typically polar

Question 34

polar molecules elute

Answer 34

SLOWER

Question 35

mobile phase is

Answer 35

typically nonpolar

Question 36

nonpolar molecules elute

Answer 36

faster

Question 37

liquid chromatography utilizes

Answer 37

silica in stationary phase toluene/nonpolar liquid used as mobile phase

Question 38

HPLC is a type of liquid chromatography that uses high pressure to pass...

Answer 38

the solvent phase through a more finely-group stationary phase increases interactions b/w molecules and stationary phase

Question 39

high-performance liquid chromatography (HPLC) has

Answer 39

higher resolving power

Question 40

gas chromatography vaporizes liquid before

Answer 40

separation

Question 41

molecules are separated based on

Answer 41

polarity and boiling point

Question 42

stationary phase in gas chromatography is a

Answer 42

thin layer of material applied in column's inside

Question 43

the polarity of the stationary phase matches that...

Answer 43

of the solute in gas chromatography

Question 44

mobile phase of gas chromatography is

Answer 44

an inert gas

Question 45

size-exclusion chromatography separates molecules by

Answer 45

size rather than polarity

Question 46

smaller molecules enter the porous gel beads, allowing them to

Answer 46

elute later

Question 47

larger molecules elute faster because

Answer 47

they do not fit in the pores and will not be slowed down

Question 48

ion-exchange chromatography separates

Answer 48

proteins by net charge

Question 49

column is filled with

Answer 49

charged beads either positive or negative part of ion exchange chromatography

Question 50

cation exchange uses

Answer 50

negative beads negative proteins elute first

Question 51

anion exchange uses

Answer 51

positive beads positive proteins elute first

Question 52

affinity chromatography separates proteins based on

Answer 52

their affinity for a specific ligand

Question 53

beads are bound to a specific ligand and proteins with a...

Answer 53

high affinity for that ligand will bind to the beads affinity chromatography

Question 54

proteins with a low affinity for the

Answer 54

ligand with elute first in affinity chromatography

Question 55

thin-layer chromatography is a

Answer 55

sheet coated in polar silica gel

Question 56

molecules are spotted at the

Answer 56

bottom of the sheet thin-layer chromatography

Question 57

sheet is placed in a nonpolar liquid and the mobile phase...

Answer 57

travels up the plate using capillary action

Question 58

nonpolar molecules have the highest

Answer 58

Rx value

Question 59

retention factor (Rf) =

Answer 59

distance spot moved / distance solvent front moved

Question 60

central dogma

Answer 60

DNA --> transcription --> RNA --> translation --> PROTEIN