Flashcards in Molecular Techniques Deck (49)
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1
What produce endonucleases
Bacteria
2
What do restriction enzymes do
Recognise and degrade foreign DNA
3
Which electrode do DNA fragments travel toward in electrophoresis and why
Positive as DNA is negatively charged
4
What sized fragments travel the furthest
Small
5
What does DNA gel electrophoresis do
Separate fragment based on size
6
How are genes cloned
Isolate specific gene with restriction enzymes and insert into plasmid vector to place into bacteria to replicate naturally
7
What are plasmids
Small circular DNA
8
Give an example of when human genes are cloned
To make human insulin
9
What enzyme concerts mRNA into cDNA
Reverse transcriptase
10
What disease is currently being treated using gene therapy
Cystic fibrosis
11
What enzyme is used in PCR and why
Taq Polymerase as it can work at higher temperatures
12
What is the first step in PCR
Heat strands to 95 degrees to denature DNA to give single strands
13
What is the second step of PCR
Cool DNA to 55 degrees to allow DNA primers to bind
14
What is the final step of PCR
Heat to 72 degrees to allows taq Polymerase to bind and add nucleotides from the 3' end.
15
Why is PCR used
Look for mutations, to see if there's a loss or gain of restriction fragments or to see size of product
16
Why can you use PCR to determine if someone has sickle cell anaemia
In sickle cell the mutation changes a restriction site and so this site will not be cut like it would in normal DNA
17
What does PCR result in
Lots of copies of a specific piece of DNA
18
What is protein gel electrophoresis
Separation of proteins
19
What different types of protein electrophoresis are there?
SDS page, isoelectric focussing and 2D page
20
What is SDS page
When proteins are denatured, given a negative charge and separate due to size as they migrate to the positive electrode
21
What is 2D page separation
Separation of proteins based on size and charge
22
What is isoelectric focussing
Separation of proteins based on charge. Protein migrate until they reach a pH equal to their pI
23
What is proteomics
Protein identification
24
How do you carry out proteomics
Digest protein, perform mass spectrometers and identify protein based on peptide sizes
25
What Techniques uses antibodies instead of DNA hybridisation
Western blotting
26
What is ELISA
A technique to detect protein concentration in a mixture by using antibodies to bind to the protein
27
What is an enzyme assay
A technique used to measure enzyme activity by measuring appearance of product or disappearance of substrate
28
The concentration of which enzyme increases during a heart attack
Creatine Kinase
29
What are monoclonal antibodies
Antibodies that can bind to 1 epitope/protein only
30
