7.1 Using Gene Sequencing

Question 1

why is a polymerase chain reaction used

Answer 1

to amplify DNA

Question 2

what does a reaction mixture contain in PCR

Answer 2

taq polymerase
DNA sample
free nucleotides
primers
buffer

Question 3

where is the reaction mixture placed

Answer 3

in a thermal cycler

Question 4

describe the process of amplifying DNA (polymerase chain reaction)

Answer 4

1. set thermal cycler at 90-95 degrees for 30 seconds so DNA strands separate
2. At 50-55 degrees for 20 seconds
   primers bind to the DNA strands- annealing
3. 72 degrees for at least a minute
   DNA polymerase builds up complementary strands of DNA

process repeats

Question 5

give four uses of DNA sequencing

Answer 5

DNA barcoding to identify species and investigate evolution
DNA sequences are shared freely between scientists
to investigate causes of disease
to manage diseases

Question 6

what does the mixture in DNA sequencing contain

Answer 6

free nucleotides
taq polymerase
primers
terminator bases- bases with slightly modified structure and fluoresces under UV light

Question 7

describe the process of DNA sequencing

Answer 7

1. 96 degrees
   separates DNA strands
2. 50 degrees
   primers bind to DNA strand
3. 96 degrees
   taq polymerase can attach free bases
4. DNA is sucked into a capillary tube containing gel
   DNA moves through gel due to negative charge
   smaller fragments move faster
5. smallest fragment passes laser beam first to identify a sequence of bases

Question 8

what do terminator bases do

Answer 8

they stop DNA taq polymerase and enzyme moves away

Question 9

what is an exon

Answer 9

part of a gene that codes for a protein

Question 10

wat is an intron

Answer 10

part of a gene that does not code for a protein

Question 11

what are introns made from

Answer 11

minisatellites and microsatellites

Question 12

what determines whether an intron is a microsatellite or minisatellite

Answer 12

the order of bases

Question 13

describe the process of gel electrophoresis

Answer 13

1. white blood cells are extracted from a sample
2. it is centrifuged and detergent added to break membrane and extract DNA
3. PCR amplifies DNA
4. specific restriction enzymes break DNA into small intron fragments
5. enzyme cuts DNA at intron sequences to give mini and microsatellites
6. DNA is negatively charged, placed in agarose gel and current is applied
7. smaller fragments move faster than larger fragments
8. creates a band of DNA
9. radioactive material is added which produces fluorescent image and copy of DNA is obtained

Question 14

how is a Southern blot carried out

Answer 14

alkaline solution
filter paper with agarose gel, nitrocellulose paper

capillary action transfers DNA from gel to nitrocellulose paper

Question 15

describe hybridisation

Answer 15

gene probes with a radioactive tag latch onto DNA so they are visible
X-rays form a permanent image of DNA bases
every individual will have a different pattern of bands