2. Studying DNA

Question 1

What at first was thought to be the carrier of genetic material and why?

Answer 1

Protein

Had greater diversity

Question 2

Griffiths Experiment

Answer 2

Smooth (bad) and rough (not bad) bacteria

Found that harmless R bacteria would be transformed by taking up genetic material from S and became harmful

Question 3

What are the bases in DNA?

Answer 3

Guanine
Adenine
Thymine
Cytosine

Question 4

What are the bases in RNA

Answer 4

Guanine
Adenine
Uracil
Cytosine

Question 5

What are the 3 main constituents of necleotides that make up DNA?

Answer 5

Base
Sugar (deoxyribose)
Phosphate

Question 6

What is Chargaff’s rule?

Answer 6

%A = %T
%G = %C

Question 7

What are the advantages of DNA over RNA?

Answer 7

DNA is more stable

Question 8

What are the key characteristics of DNA?

Answer 8

Capacity for replication
Capacity to store information
Capacity to express information
Capacity for variation by mutation

Question 9

What is the structural difference between RNA and DNA’s sugar?

Answer 9

Ribose has an O molecule on the bottom of the 2’C

Deoxyribose doesn’t

Question 10

What bond holds the DNA backbone together?

Answer 10

3’-5’ phosphodiester bonds

Question 11

Which of the bases are purines?

Answer 11

Adenine

Guanine

Question 12

Which of the bases are pyrimidines?

Answer 12

Uracil
Thymine
Cytosine

Question 13

Why are H bonds good for joining strands of DNA?

Answer 13

Week enough to be seperated in localised areas without using too much energy
Strong enough overall to hold it together

Question 14

What’s at the 5’ end of DNA?

Answer 14

A phosphate group

Question 15

What’s at the 3’ end of DNA

Answer 15

A hydroxyl group

Question 16

Which part of DNA is hydrophlic?

Answer 16

The sugar-phosphate backbone

Question 17

Which part of DNA is hydrophobic?

Answer 17

The bases

Question 18

What is the general method for extracting DNA?

Answer 18

Sample disruption
Release of cell contents
Extraction
Purify and concentrate DNA

Question 19

What methods of disrupting samples to get DNA are there?

Answer 19

Mechanical
Chemical lysis
Osmotic - put them in pure water and they burst due to osmotic shock
Enzymatic

Question 20

Lysate

Answer 20

The contents of a lysed cell

Question 21

Traditional method of extracting DNA

Answer 21

Mix cells with phenol-chloroform solution and centrifuge
Forms solvent and aqueous phase
DNA in aqueous phase

Question 22

Why does DNA appear in the aqueous phase in phenol-chloroform extraction?

Answer 22

Because DNA is hydrophilic

Question 23

Ethanol precipitation method of purify and concentrate DNA

Answer 23

DNA is insoluble in ethanol

High speed centrifugation makes a pellet of DNA that can be re-suspended in a small volume of clean H2O

Question 24

What are the advantages of nucleic acid purification kits?

Answer 24

Allow preparations of high purity

Quick and easy

Question 25

What are the disadvantages of nucleic acid purification kits?

Answer 25

Expensive | Solutions may be of unknown compisition

Question 26

What setting on the spectophotometer reads how much DNA is in a sample?

Answer 26

A260

Question 27

What setting on the spectophotometer reads the purity of DNA in a sample?

Answer 27

A280

Question 28

What is the optical density to dsDNA (ng/ul) equivalent?

Answer 28

1.0 OD = 50ng/ul

Question 29

Equation for the concentration of DNA using a spectrophotometer?

Answer 29

DNA conc (ng/ul) = A260 OD x dilution factor x 50

Question 30

Equation for the purity of DNA using a spectrophotometer?

Answer 30

= A260 / A280

Question 31

What does a ratio of A260 and A280 at ~1.8 tell us the sample is?

Answer 31

Pure DNA

Question 32

What does a ratio of A260 and A280 at ~2.0 tell us the sample is?

Answer 32

Pure RNA

Question 33

What does a ratio of A260 and A280 at <1.6 tell us the sample is?

Answer 33

Other contaminants

Question 34

Why do A-T base pairs separate at a lower temperature than C-G pairs?

Answer 34

A-T has 1 less H bond

Question 35

What does the Tm value indicate when heating DNA?

Answer 35

The temperature at which half the base pairs are denatured and half remain intact

Question 36

If two different samples have different Tm values what does this tell us?

Answer 36

The DNA with the higher Tm value has a greater amount of C-G base pairs

Question 37

What is reannealing of DNA?

Answer 37

When single strands of DNA find their complimentary strands to reform a double helix

Question 38

What happens to DNA if you cool it quickly after melting it?

Answer 38

The single strands don't reanneal

Question 39

Non-repetitive DNA

Answer 39

Sequences that are unique and occur in only one place in the genome

Question 40

Repetitive DNA

Answer 40

DNA sequences that are present in more than one copy per genome

Question 41

Multigene families

Answer 41

Collection of similar genes | Often clustered together

Question 42

What is a pseugogene?

Answer 42

Genes that have lost protein coding ability

Question 43

What can cause pseudogenes?

Answer 43

``` A premature stop codon Insertion Deletion Frameshift mutation Loss of a promotor ```

Question 44

Highly repetitive DNA (satellite DNA)

Answer 44

Short sequences repeated many times

Question 45

Types of middle repettitive DNA

Answer 45

Minisatellites LINES SINES

Question 46

Minisatellites

Answer 46

Variable number of tandem repeats Sequences about 5-100 bp long Repeated many times end to end Varies between individuals

Question 47

Which type of repetitive DNA is used in DNA fingerprinting?

Answer 47

Minisatellites

Question 48

LINES

Answer 48

Long interspersed sequences | Up to 6000 bp

Question 49

SINES

Answer 49

Short interspersed sequences 100-500 bp Don't code for anything

Question 50

Transposons

Answer 50

Jumping genes | Piece of DNA that can move around then genome

Question 51

What is the cause of repetitive DNA?

Answer 51

Transposons

Question 52

What enzymes cut DNA?

Answer 52

Restriction enzymes

Question 53

Why are blunt ends harder to allow annealing than sticky ends?

Answer 53

No H bonds form from overlapping tails of DNA | Makes it harder for DNA ligase to make the phosphodiester bonds

Question 54

Is DNA positively or negatively charged?

Answer 54

Negatively

Question 55

How does gel electrophoresis work?

Answer 55

Negative charged DNA moves towards + electrode | Smaller DNA can fit through gaps in gel better than large so travel closer to the + electrode

Question 56

How does DNA move in gel electrophoresis?

Answer 56

By reptation

Question 57

What do you use to visualise the DNA in gel elctrophoresis?

Answer 57

Ethidium bromide

Question 58

What tool is used to compare the movement of the DNA samples in gel electrophoresis?

Answer 58

A DNA ladder

Question 59

Polyacrylamide gels

Answer 59

Better for smaller fragments of DNA than agarose | Much better resolution - can distinguish difference of 1 bp

Question 60

How do plasmids behave in agarose?

Answer 60

They can supercoil under the tortional stress | Supercoiled plasmids move move than non-supercoiled as they are more streamlined

Question 61

Restriction fragment length polymorphisms (RFLP)

Answer 61

Can be used to detect some diseases caused by a single base pair change

Question 62

How can RFLP be used be used to detect diseases caused by a single base pair change?

Answer 62

Single change causes a loss of a a particular recognition site for an restriction enzyme When applied the restriction enzyme doesn't make the cut it usually would in that place Using gel electrophoresis you can identify the longer than normal length of DNA that wasn't cut

Question 63

Name 3 uses of restriction enzymes

Answer 63

Cutting and pasting genes for cloning Checking whether cloning was successful Diagnostics

Question 64

What is the hyperchromic effect?

Answer 64

DNA when heated becomes single stranded ssDNA is less compact than dsDNA ssDNA will have a higher A260 reading

Question 65

Which base pairs separate at a lower temperature? | Why?

Answer 65

A-T | They have 2 H bonds between them while C-G has 3

Question 66

How is aragose gel made?

Answer 66

Boiled with water then cooled

Question 67

What are tandem repeats?

Answer 67

Repeated sequences of DNA or RNA that lie next to each other

Question 68

Satellite DNA

Answer 68

Highly repetitive DNA Found as clusters of tandem repeats Permanently coiled tightly into heterchromatin

Question 69

What does VNTR stand for?

Answer 69

Variable number tandem repeats

Question 70

Variable number tandem repeats

Answer 70

Short tandem repeats | Fewer copies than satellite DNA