Molecular Diagnostics

Question 1

What is the probe in Southern Blotting?

Answer 1

DNA
SNOW
DR P

Question 2

What is the target in Southern Blotting?

Answer 2

DNA

both are DNA

Question 3

What is the target in Northern Blotting?

Answer 3

mRNA

Snow drop

Question 4

What is the probe in Northern Blotting?

Answer 4

single stranded DNA

Question 5

What is the target in western blotting?

Answer 5

Protein

Question 6

What is the basic sequence of PCR?

Answer 6

put in your dna, primers, and enzymes
denature DNA by heating
cool it down to allow primers to anneal
Add dNTPs
Taq pol will extend primers

Question 7

What is the disadvantage of PCR?

Answer 7

need to know the sequence you are looking to amplify

Question 8

What is the point of PCR?

Answer 8

to amplify a sequence or see if it is there at all

Question 9

What does the number of cycles mean in qPCR?

Answer 9

high number of cycles = little protein

small number of cycles = a lot of protein

Question 10

What is restriction fragment length polymorphism (RFLP) and what is it used for?

Answer 10

Detects differences in genomes by comparing the lengths of fragments after dna is cut with endonucleases
Used for forensics, paternity testing and disease detection

Question 11

How many restriction sites does a normal beta globulin allele have?

Answer 11

3 –> creates 2 pieces of gene = 2 bands on normal blot

Question 12

How many restriction sites does a sickle cell beta globulin allele have?

Answer 12

2 –> only one piece of dna = 1 band on homozygous pt; 3 on heterozygous (1 copy of normal, 1 copy of abnormal)

Question 13

What do you use variable number of tandem repeats (VNTR) for?

Answer 13

identification and severity of inherited diseases

Huntington’s

Question 14

What is a plasmid vector?

Answer 14

Circle of DNA from bacteria; where you put your human gene to produce something

Question 15

What is recombinant dna?

Answer 15

Plasmid vector with human dna in it

Question 16

What type of cells are taken from an animal to produce monoclonal antibodies?

Answer 16

spleen cells

Question 17

What are spleen cells fused with to make immortal cells for monoclonal antibody production?

Answer 17

myeloma cells –> hybridoma

Question 18

What does an indirect ELISA measure?

Answer 18

a specific antibody

antigen coated well –> Ab binds –> secondary Ab w/ enzyme binds –> color change

Question 19

What does a sandwich ELISA measure?

Answer 19

A specific protein/antigen

Antibody coated wells –> Antigen binds –> secondary Ab w/ enzyme binds –> color change

Question 20

What technique is used to first detect HIV?

What type of molecule is it testing for?

Answer 20

ELISA

Testing for antibodies produced 4-6 weeks after infection, so use indirect ELISA

Question 21

What technique is used to detect an MI?

Answer 21

looking for troponin –> Use sandwich ELISA bc looking for an antigen

Question 22

Which type of ELISA is used in pregnancy tests?

Answer 22

looking for an antigen (hCG) –> use sandwich ELISA

Question 23

What technique is used in the confirmation of HIV?

Answer 23

Western blotting

Question 24

What is the general process of western blotting?

Answer 24

SDS-PAGE to separate proteins –> transfer to nitrocellulose membrane –> and 1 antibody –> and 2 antibody w/ enzyme tag –> and substrate –> color that is proportional to amount of protein