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Biology 2 > Lecture 16 DNA Hereditary > Flashcards

Lecture 16 DNA Hereditary Flashcards

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1
Q

How did dye that binds to DNA and turned it red provide circumstantial evidence that DNA was the genetic material?

A
  • It was in the right place
  • It varied among species
  • It was present in the right amounts
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2
Q

In what ways was DNA present in the right amounts in order to be the genetic material?

A

DNA is somatic cells was twice that of reproductive cells

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3
Q

On what organism was Frederick Griffith studying in the 1920’s?

A

Streptococcus pneumoniae (causes pneumonia)

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4
Q

What two strains of pneumonia was Griffith working with?

A

S strain and R strain

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5
Q

What did cells of the S strain of S. pneumoniae produce?

A

Colonies that looked smooth and was covered by polysaccharide capsule

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6
Q

What is the significance of the polysaccharide capsule covering S. pnuemoniae?

A

They are protected from attack by the host cells immune system

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7
Q

What happened when S strain was injected into mice?

A

Caused pneumonia- strain was virulent

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8
Q

What did the cells of the R strain of S. pneumoniae produce?

A

Colonies that looked rough (lacked capsule, not virulent)

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9
Q

What did Griffith do to some mice?

A

Inoculate them with heat-killed S. pneumoncocci (heat killed bacteria does not cause infection)

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10
Q

What happened when Griffith inoculated some mice with a mixture of living R bacteria and heat killed S bacteria?

A

The mice died of pneumonia

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11
Q

What happened when Griffith examined the blood of mice that died after being inoculated with a mixture of living R bacteria and heat killed S bacteria?

A

Full of living bacteria, many with S strain characteristics

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12
Q

What was Griffith’s conclusion?

A

A chemical substance from one cell is capable of transforming another cell

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13
Q

At the time, what was the chemical that could cause a heritable change in affected R cells called?

A

a chemical transforming principle

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14
Q

Who identified the transforming principle?

A

Oswald Avery and his colleagues

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15
Q

What did Oswald Avery and his colleagues do to identify the transforming principle as DNA?

A

They treated samples known to contain pnemococcal transforming principle in a variety of ways to destroy different types of molecules (proteins, nucleic acids, carbohydrates, lipids) and tested samples to see if they had retained transforming activity

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16
Q

What did Oswald Avery and his colleagues find in their experiments?

A

If DNA was destroyed, transforming activity was lost, but there was no loss of activity when other molecules were destroyed

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17
Q

What was the final step taken by Oswald Avery and Colin Macleod and Macyln McCarty in their experiment?

A

To isolate virtually pure DNA from the sample containing pnuemococcal transfomring principle and showed that it caused bacterial transformation

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18
Q

Why did the work done by Avery, MacLeod and McCarty have little impact when it was published?

A

Scientists did not believe DNA was chemically complex enough to be genetic material
Bacterial genetics was new- it was not clear bacteria even had genes

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19
Q

What enzymes were used in Avery’s experiments?

A

RNase
Protease
DNase

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20
Q

What did the Hershey-Chase experiment sought to determine?

A

Whether DNA or protein was genetic material

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21
Q

What was the Hershey-Chase experiment carried out with?

A

Bacteriophage T7- consisting of DNA inside a protein coat

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22
Q

What did Hershey and Chase deduce about the entry of some viral components?

A

Entry of some viral components affects genetic program of the host bacterial cell, transforming it into a bacteriophage factory

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23
Q

How did Hershey and Chase trace the components of the bacteriophage over its life time?

A

Radioactive isotopes

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24
Q

What radioactive isotope is used to mark protein?

A

Sulfur 35

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25
Q

Why is sulfur 35 used to mark protein?

A

Sulfur is in the amino acid cysteine and methionine

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26
Q

How did Hershey and Chase label the bacteriophage with sulfur 35?

A

They grew the bacteriophages in the presence of S35

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27
Q

What element is rich in DNA and where?

A

Phosphorus

In the deoxyribose-phosphate backbone

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28
Q

What radioisotope is used to mark DNA?

A

Phosphorus 32

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29
Q

What method did Hershey and Chase use to test their hypothesis?

A

32-P labelled bacteriophage infects bacteria in one experiment, 35-S labelled bacteriphage infects bacteria in the other

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30
Q

What happened after the two different experiments were infected with labelled bacteriophages?

A

After a few minutes, the mixture was agitated in a blender without bursting bacteria to strip away parts of the bacteriophage that had not penetrated the bacteria

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31
Q

What happened after blending the mixture in the Hershey-Chase experiment?

A

The centrifuged it to separate the bacteria from the rest of the material

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32
Q

What did the scientists find after centrifuging the experiments?

A

Supernatant contained mostly sulfur (and thus viral protein) and phosphorus had remained mostly with the bacteria in the pellet

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33
Q

What did the distribution of sulfur and phosphorus in the centrifuged supernatant/pellet suggest?

A

DNA had been transferred to bacteria, thus DNA was the compound responsible for redirecting genetic program of bacterial cell

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34
Q

What other experiments did Hershey and Chase perform?

A

Longer range experiments in which progeny viruses were collected- almost none contained labelled sulfur, but about 1/3rd had original phosphorus

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35
Q

What was the logical conclusion to Hershey and chase’s longer range experiment?

A

Because DNA was carried over in the virus from generation to generations, protein was not, the hereditary information of the virus is contained in the DNA

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36
Q

What is genetic transformation of eukaryotic cells by DNA called?

A

Transfection

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37
Q

How is transfection demonstrated?

A

Using a marker

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38
Q

What is a marker?

A

A gene whose presence in the recipient cell confers an observable phenotype

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39
Q

What marker gene did researchers use to demonstrate transfection in eukaryotic cells?

A

Nutritional or antibiotic resistance marker genes that permit the growth of transformed recipient cells but not nontransformed cells

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40
Q

What is thymidine kinase?

A

An enzyme needed to make use of thymidine

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41
Q

What happens in the absence of the gene that codes for thymidine kinase?

A

Mammalian cells do not grow

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42
Q

What happens when DNA containing thymidine kinase marker gene is added to a culture of mammalian cells? What does this show?

A

Some cells will grow, demonstrating they have been transfected with the gene

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43
Q

What is a whole new genetically transformed organism called?

A

A transgenic organism

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44
Q

What was crucial evidence for deciphering the structure of DNA?

A

X-ray crystalography

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45
Q

What X-ray crystalography?

A

Isolated chemical substances that form crystals, positions of atoms can be inferred from pattern of diffraction passing through

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46
Q

What shape did Rosalind Franklin propose based on X-ray crystalography evidence?

A

Spiral or helical molecule

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47
Q

Who reported that A=T and C=G?

A

Erwin Chargaff (created Chargaff’s rule)

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48
Q

How was the solution to the puzzle of the structure of DNA accelerated?

A

Model building

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49
Q

What does model building entail?

A

Assembly of 3D representations of possible molecular structures using known relative molecular dimensions and bond angles

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50
Q

Who used the model building technique to build a single, coherent DNA molecule?

A

Watson and Crick

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51
Q

What conclusions did Watson and crick use to build their model of DNA?

A

DNA is helical

DNA is antiparallel

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52
Q

What are the 4 key features that define DNA structure?

A
  • Double stranded helix of uniform diameter
  • Right handed
  • Antiparallel
  • Outer edges of the nitrogenous bases are exposed in the major and minor grooves
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53
Q

How are the two chains of DNA held together?

A

Hydrogen bonding between specifically paired based

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54
Q

How many hydrogen bonds form between adenine and thymine?

A

2

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55
Q

How many hydrogen bonds form between cytosine and guanine?

A

3

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56
Q

What is the pattern of pyrimidines pairing with purines known as?

A

Complementary base pairing

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57
Q

What does it mean that DNA molecules are antiparallel?

A

The 3 ‘ end pairs with the 5’ end of the other strand and vice versa

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58
Q

What chemical group is at the 5’ end?

A

OPO3-

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59
Q

What chemical group is at the 3’ end?

A

OH

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60
Q

What is the significance of base exposure in grooves?

A

Hydrogen bonded base pairs are accessible for potential hydrogen bonding

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61
Q

How are the surfaces of the AT and GC base pairs chemically distinct surfaces?

A

C=O group in T and N group in A are places for additional hydrogen bonding
Additional hydrogen bonding opportunities in GC

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62
Q

What is the significance of access to exposed base pairs in major and minor grooves?

A

Key to protein-DNA interactions in replication and expression

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63
Q

What are the 4 functions of DNA?

A
  • Store genetic information
  • Susceptible to mutation
  • Precise replication
  • Expression as the phenotype
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64
Q

What three substances did Arthur Kornberg show as needed to produce DNA with the same base composition as parental DNA in a test tube?

A
  • Substrates deoxyribonucleoside triphosphates dATP, dCTP, dGTP and dTTP
  • DNA polymerase enzyme
  • DNA (template)
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65
Q

What were the three possible replication patterns that could occur in the experiment?

A
  • Semiconservative replication
  • Conservative replication
  • Dispersive replication
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66
Q

What is semiconservative replication?

A

Each parent strand serves as a a a template for a new strand, two new DNA molecules each have one old and one new strand

67
Q

What is conservative replication?

A

The original double helix serves as a template for, but does not contribute to, a new double helix

68
Q

What is dispersive replication?

A

Fragments of the original DNA molecule serves as a template for assembling two new molecules, each containing old and new parts, perhaps at random

69
Q

What type of replication did Watson and Crick’s original paper suggest?

A

Semiconservative replication

70
Q

Who demonstrated that replication is semiconservative?

A

Meselson and Stahl

71
Q

How did Meselson and Stahl distinguish old parent strands of DNA from newly copied ones?

A

Density labelling

72
Q

What isotope did Meselson and Stahl use in their experiments do distinguish parent and copied DNA strands?

A

Heavy nitrogen (nitrogen-15): nonradioactive, heavy isotope

73
Q

What bacterium did Meselson, Stahl and Jerome use in their experiments?

A

Escherichia coli

74
Q

What two cultures of Escherichia coli did they grow?

A

One in nitrogen source was heavy Nitrogen

One in which nitrogen sources which was nitrogen 14 (light)

75
Q

What was the nitrogen medium used in Meselson and Stahl’s experiments?

A

Ammonium chloride

76
Q

How was the weight of the cultures determined?

A

Combined and centrifuged cultures, DNA samples were distinguished by different densities

77
Q

How often did their bacteria cultures divide under the conditions used?

A

20 minutes

78
Q

What happened when researchers transferred N-15 E.coli to normal N-14 medium and allowed it to continue growing?

A

They collected bacteria after each division

They extracted DNA from samples

79
Q

What were the different density gradients found in each generation of the E.coli populations?

A
  • Dense after transfer (uniformly N-15)
  • After one generation, all DNA was intermediate density
  • After two generations, two equally large DNA bands: one low density and one intermediate density
80
Q

What was the density gradient in subsequent generations?

A

Proportion of low-density DNA increased steadily

81
Q

What are the two steps of DNA replication?

A
  • DNA double helix unwinds to separate template strands

- New nucleotides are joined by phosphodiester linkages to each new growing strand in sequence

82
Q

To which end are nucleotides added to the growing new strand?

A

The 3 prime end (free -OH group)

83
Q

What is the name of the huge protein complex the template strand interacts with to replicate DNA?

A

The replication complex

84
Q

What is the purpose of the replication complex?

A

To catalyze the reactions involved

85
Q

To what does the replication complex initially bind?

A

A base sequence called the origin of replication (ori)

86
Q

In what ways does DNA replicate from the origin of replication?

A

In both directions, forming two replication forks

87
Q

What is the binding of the replication complex to the ori a result of?

A

Recognition of different nucleotide bases by proteins

88
Q

What part moves and what part remains stationary during DNA replication?

A

The replication complex remains stationary, DNA threads through

89
Q

How does DNA enter and exit the replication complex?

A

Enters as a single strand

Exits as double strands

90
Q

What do all replication complexes contain?

A

Several proteins with different roles in DNA replication

91
Q

What is the first event at the origin of replication?

A

Localized unwinding (denaturation) of DNA

92
Q

What forces hold the two strands together?

A
  • Hydrogen bonding

- Hydrophobic interactions of the bases

93
Q

What enzyme causes the DNA to unwind?

A

DNA helicase

94
Q

What does DNA helicase use to make DNA unwind?

A

Energy from ATP hydrolysis

95
Q

What binds to unwound DNA to keep unwound strands from re-associating?

A

Single-strand binding proteins

96
Q

What is the purpose of unwinding DNA?

A

Makes the two template strands available for complementary base pairing

97
Q

How many base pairs are there in small circular chromosomes such as those found in bacteria?

A

1-4 million

98
Q

Where do small circular chromosomes replicate from?

A

A single origin

99
Q

What happens as DNA from a small circular chromosome moves through the replication complex?

A

The replication forks grow around the circle, two interlocking circular DNAs are formed

100
Q

How are the two interlocked DNA’s separated?

A

using enzyme DNA topoisomerase

101
Q

How fast are DNA polymerases in E.coli?

A

1000 bases per second, 20-40 minutes for replication of all base pairs

102
Q

How fast are DNA polymerases in humans?

A

50 bases per second- finished in an hour due to many polymerases working at many replication forks

103
Q

Where are the origins of replication in large, linear chromosomes (such as human chromosomes)?

A

Many places

Adjacent ori’s can be bound at the same time

104
Q

What shape is DNA polymerase?

A

Open right hand with a palm, thumb and fingers

105
Q

How do DNA polymerases elongate polynucleotide strands?

A

By covalently linking new nucleotides to a previously existing strand

106
Q

What is needed for DNA polymerase to being adding nucleotides to a chain?

A

A primer- a short single stand of RNA

107
Q

What is the primer complementary to?

A

The DNA template strand

108
Q

What synthesizes the primer?

A

An enzyme called primase

109
Q

What do the ‘fingers’ of DNA polymerase do?

A

Recognize different shapes of the 4 nucleotide bases

110
Q

How does primase work?

A

It binds to the template strand and synthesizes the primer, primase is released, DNA polymerase binds

111
Q

What happens once DNA replication has been completed?

A

The primer is degraded
DNA is added in its place
Resulting DNA fragments are connected by enzyme action

112
Q

What DNA polymerases are there in cells?

A

One is responsible for chromosomal DNA replication

The others are involved in primer removal and DNA repair

113
Q

How many DNA polymerases have been identified in humans?

A

14,

114
Q

Which DNA polymerase catalyzes replication in humans?

A

DNA polymerase 𝛿

115
Q

What DNA polymerase catalyses replication in E.coli? How many are there?

A

DNA polymerase III

5

116
Q

What name is given to the newly replicating strand?

A

The leading strand

117
Q

What direction is the leading strand pointing in?

A

The right direction to grow continuously at its 3’ end

118
Q

What name is given to the other strand (not the leading strand)

A

The lagging strand

119
Q

What direction is the lagging strand pointing in?

A

The wrong direction- its 3’ end gets further and further from the fork

120
Q

How is the lagging strand synthesized?

A

Small, discontinous stretches (100-200 nucleotides at a time, 1000-2000 in prokaryotes) from 5’ to 3’ end

121
Q

What are the small, discontinous stretches of DNA on the lagging strand called?

A

Okazaki fragments

122
Q

What does each Okazaki fragment require?

A

Its own primer

123
Q

In bacteria, how does DNA polymerase III synthesize Okazaki fragments?

A

By adding nucleotides to a primer until it reaches the primer of the previous fragment

124
Q

What happens when DNA polymerase III reaches the primer of the previous fragment?

A

DNA polymerase I removes the old primer and replaces it with DNA

125
Q

What enzyme catalyzes the formation of the phosphodiester linkage between the Okazaki fragments after the primer is removed?

A

DNA ligase

126
Q

What are DNA polymerases that makes them so fast?

A

Processive

127
Q

What does it mean that DNA polymerases are processive?

A

They catalyze many polymerizations each time they bind to a DNA molecule

128
Q

How is the newly replicated strand stabilized?

A

Using a sliding DNA clamp

129
Q

What is the structure of a sliding DNA clamp?

A

Multiple identical subunits assembled into a doughnut shape

130
Q

What does the DNA clamp do?

A

Binds to DNA just behind DNA polymerase, keeping it tightly associated with newly replicated DNA

131
Q

What happens if the DNA clamp is absent?

A

DNA polymerase dissociates from DNA after 20-100 polymerizations

132
Q

How many polymerizations can occur when the DNA clamp is present?

A

50,000

133
Q

Why are telomeres not replicated?

A

When terminal primer of Okazaki fragments is removed, no DNA can be synthesized to replace it. New chromosome has a bit of single stranded DNA at each end

134
Q

What happens when each new chromosome has a bit of single stranded DNA at each end?

A

Mechanisms cut off single stranded region along with some intact double stranded end

135
Q

What do many eukaryotes have at the end of their chromosomes?

A

Repetitive sequences at the ends of chromosomes called telomeres

136
Q

What is the telomere sequence in humans?

A

TTAGGG, repeated 2,500 times

137
Q

What happens after 20-30 divisions in humans?

A

The chromosomes are unable to take part in cell division and the cell dies

138
Q

How do constantly dividing cells such as bone marrow stem cells and gamete producing cells maintain telomeric DNA?

A

An enzyme, telomerase, catalyzes addition of lost telomeric sequences

139
Q

Where is telomerase also expressed?

A

90% human cancers

140
Q

What is the observed error rate in base replications in humans?

A

1 in 10^5 bases, 60,000 mutations for every human cell division

141
Q

What are the three main DNA repair mechanisms that cells have?

A
  • Proofreading mechanism
  • Mismatch repair
  • Excision repair
142
Q

How does DNA polymerase perform a proofreading function?

A

It recognizes mispairing bases and removes the nucleotide and tries again
Lowers error rate to 1 in 10^10

143
Q

What happens to repair DNA after DNA has been replicated?

A

A second set of proteins surveys the newly replicated molecule and looks for mismatches

144
Q

How does mismatch repair mechanism detect wrong bases?

A

DNA strand is chemically modified after replication, methyl groups are added to adenines in prokaryotes. Unmethylated strands are therefore ‘marked’ and errors should be repaired

145
Q

How is damage to DNA molecules repaired?

A

Excision repair

146
Q

What does excision repair do?

A

Removes abnormal bases that have formed because of chemical damage and replaces them with functional groups

147
Q

What is the name of the reaction that makes multiple copies of DNA?

A

The polymerase chain reaction

148
Q

PCR reaction is_____, the sequence of steps is repeated over and over again.

A

Cyclic

149
Q

What are the three steps of the PCR reaction?

A
  • Double stranded fragments of DNA are separated into single strands by heating
  • Short, artificial primer is added along with 4deoxyribonucleotide triphospates and DNA polymerase
  • DNA polymerase catalyzes production of complementary strands
150
Q

What is the exponential increase in the number of copies of DNA sequence called?

A

Amplifying sequence

151
Q

What does the PCR technique require?

A

That the base sequences at the 3’ end are known so primer can be made

152
Q

What bacterium lives in the hot springs of Yellowstone National park?

A

Thermus aquaticus

153
Q

What temperature does Thermus aquaticus live up to?

A

95 degrees celcius

154
Q

What technique allows researchers to determine base sequences?

A

DNA sequencing

155
Q

What does DNA sequencing rely on?

A

Artificially altered nucleotides

156
Q

What sugar replaces deoxyribose in nucleotide sequencing?

A

2,3-dioxyribose

157
Q

What is the result of replacing deoxyribose with 2,3-dioxyribose?

A

dideoxyribonucleoside triphosphate

158
Q

What happens when dideoxyribonucleoside triphosphate is added to the polynucleotide chain?

A

Lacks -OH group at 3’, next nucleotide is not added, synthesis stops

159
Q

What are the steps of DNA sequencing?

A
  1. Denature

2. Mix with DNA polymerase, synthesized primers, 4 dNTP’s and ddNTP’s bonded to fluorescent tags

160
Q

What happens after mixing denatured DNA to be sequenced with DNA polymerase, synthesized primers, 4 dNTP’s and ddNTP’s bonded to fluorescent tags?

A

DNA replication proceeds, test tube contains varying length synthesized strands, fragments are denatured from templates

161
Q

What happens to the fragments that were denatured from templates?

A

Subject to electrophoresis

162
Q

What does electrophoresis do?

A

Separates DNA fragments by length and detect differences as short as one base

163
Q

What happens during electrophoresis?

A

Laser beam excites fluorescent tags- colour is detected and recorded. Length is fed into computer which prints DNA sequence of the fragment.

Biology 2 (14 decks)

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