blotting

Question 1

blotting is the transfer of?

Answer 1

DNA, RNA, or proteins from an electrophoresis gel to a nitrocellulose of PVDF membrane so that further experiments can isolate particular nucleic acid fragment or protein (probing)

Question 2

southern blotting is the detection of specific?

Answer 2

DNA sequence in a sample

Question 3

northern blotting is the detection of specific?

Answer 3

RNA sequence in a sample

Question 4

in southern blotting, the first step is?

Answer 4

the DNA strand of interest is exposed to restriction enzymes that cut the DNA strand into smaller fragments

Question 5

in southern blotting, the second step is?

Answer 5

the new cleave DNA are denatured using NaOH to create ssDNA strands

Question 6

in southern blotting, the third step is?

Answer 6

the ssDNA (single stranded DNA) undergoes gel electrophoresis to separate them based on size
with the smaller fragment at the bottom

Question 7

in southern blotting, the fourth step is?

Answer 7

the DNA from the gel is transferred to a sheet of nitrocellulose paper and exposed to a 32P radio-labeled DNA probe that is complementary to DNA of interest

Question 8

the probe is constructed with radiolabeled nucleotides in order to?

Answer 8

visualize the target sequence with special film

Question 9

in southern blotting, the fifth step is?

Answer 9

the nitrocellulose paper is viewed using autoradiography to identify the strand of interest

Question 10

in northern blotting, the steps are the same as southern blotting (except steps 1 and 2) just using?

Answer 10

RNA which allows you to determine whether specific gene products are normal or pathologic

Question 11

in both southern and northern blotting, polyacrylamide is used if the strands are?

Answer 11

less than 500 base pairs

Question 12

in both southern and northern blotting, agarose is used if the strands are?

Answer 12

more than 500 base pairs

Question 13

western blotting allows for the detection of?

Answer 13

specific proteins in a sample

Question 14

western blotting is used in real life to determine whether?

Answer 14

cancer cells express certain tumor-promoting growth receptors on their surface

Question 15

in western blotting, the first step is?

Answer 15

cells are collected and solubilized in detergent to release their cytoplasmic contents

Question 16

in western blotting, the second step is?

Answer 16

cell lysates are denatured, so they can be load next to the later
(cell lysates contain hundreds of different proteins)

Question 17

in western blotting, the third step is?

Answer 17

protein sample is loaded into SDS-PAGE gel and separated based on size
the proteins are negatively charged so that the smallest will migrate farthest from the wells

Question 18

in western blotting, the fourth step is?

Answer 18

proteins from gel are transferred to polymer sheet and exposed to radio-labeled antibody probe that is specific to protein of interest

Question 19

in western blotting, the fifth step is?

Answer 19

the polymer sheet is viewed using autoradiography and the protein of interest is bound to the radio-labeled antibody will be visible