Ch6: DNA and Biotech

0.0(0)
studied byStudied by 0 people
learnLearn
examPractice Test
spaced repetitionSpaced Repetition
heart puzzleMatch
flashcardsFlashcards
Card Sorting

1/102

encourage image

There's no tags or description

Looks like no tags are added yet.

Study Analytics
Name
Mastery
Learn
Test
Matching
Spaced

No study sessions yet.

103 Terms

1
New cards
Deoxyribonucleic Acid (DNA)
Store genetic info

Polydeoxyribonucleotide from linked monodeoxyribonucleotides

In chromosomes, mitochondria, and chloroplasts
2
New cards
Nucleosides
Pentose sugar (C-1’) + nitrogenous base
3
New cards
Nucleotides
Nucleoside (C-5’) + phosphate groups

Named for # of phosphate groups

DNA building blocks
4
New cards
Nucleotides: ADP and ATP
High energy from repulsion of - phosphate groups

Bond breaking = exothermic
5
New cards
Nucleic Acid Classification
Pentose sugar

Ribose: RNA

Deoxyribose: DNA
6
New cards
Sugar-Phosphate Backbone
Read 5’ to 3’

Form from nucleotides joined by phosphodiester bonds 3’-5’ (3’ C to 5’ phosphate)

Overall neg charge
7
New cards
DNA Directionality
Polarity from 5’ and 3’

5’: -OH or phosphate bonded to sugar C-5’

3’: -OH on sugar C-3’
8
New cards
Reading DNA
5’ to 3’: 5’-ATG-3’

3’ to 5’: 3’-GTA-5’

Phosphates: pApTpG

Deoxyribose: dAdTdG
9
New cards
Nitrogenous Base: Purines
**PURE AGriculture**

2 rings

Adenine (A) and guanine (G)

In DNA and RNA
10
New cards
Nitrogenous Base: Pyrimidines
**PYRAMIDS can CUT**

1 ring

Cytosine (C), thymine (T), uracil (U)

In DNA (C and T) and RNA (U)
11
New cards
Aromatic
Stable ring (From delocalized π e)


1. Cyclic compound
2. Planar compound
3. Conjugated (alternate single and double bonds)
4. 2n+2 π e (Huckel’s rule)
12
New cards
Heterocycles
Ring structures with 2+ different elements in ring
13
New cards
Aromatic Heterocycles
Nitrogenous base structure give nucleic acids stability
14
New cards
Watson-Crick DNA Model
Antiparallel double helix

Sugar-phosphate backbone outside, nitrogenous bases inside

Complementary base-pairing
15
New cards
Complementary Base-Pairings
A and T/U: 2 H bonds

G and C: 3 H bonds (stronger)
16
New cards
Chargaff’s Rule
In DNA:

%A = %T

%C = %G

%purines = %pyrimidines
17
New cards
B-DNA
Right-handed DNA helix

Grooves between strands (protein binding sites)
18
New cards
Z-DNA
Left-handed zigzag DNA helix

From high GC-content or high salt concentration

No biological activity
19
New cards
DNA Denaturation
Disrupt H bonds and base-pairings

Intact covalent bonds

Separate single DNA strands from helix
20
New cards
Denaturation Agents
Heat, alkaline pH, chemicals (formaldehyde and urea)
21
New cards
DNA Reannealing
Repairing single-stranded DNA into helix

Remove denaturing conditions
22
New cards
Nucleoproteins
Proteins associating with DNA

Acid soluble and stimulate transcription
23
New cards
Histones
Nucleoprotein (basic)

Wind DNA into chromatin

5 proteins (H1, H2A, H2B, H3, H4)
24
New cards
Nucleosome
Histone core with 2 protein copies (H2A, H2B, H3, H4)

200 DNA base pairs wrapped around
25
New cards
H1 Protein
Seal DNA entering and leaving nucleosome

Increase stability
26
New cards
Chromatin
DNA associated with histones
27
New cards
Heterochromatin
Condensed chromatin around histones (small amount)

Repetitive DNA sequences

Dark under microscope

Transcriptionally silent
28
New cards
Euchromatin
Uncondensed chromatin unbound from histones

Light under microscope

Genetically active
29
New cards
Telomere
Repeating TTAGGG end sequence

Lost during replication to prevent info loss

Prevent unraveling (high GC-content)
30
New cards
Telomerase
Replace lost telomeres
31
New cards
Centromere
Chromosome centre

Site of construction

Contain heterochromatin with high GC-content (connect sister chromatids until separated by microtubules)
32
New cards
Replication: Replisome/Replication Complex
Specialized proteins assisting DNA polymerase
33
New cards
Replication: Origin of Replication
DNA unwinding site

Replication forks form on either side
34
New cards
Origin of Replication: Prokaryotes
1 origin

Produce 2 identical circular DNA molecules
35
New cards
Origin of Replication: Eukaryotes
Multiple origins

Produce sister chromatids connected at centromere
36
New cards
Replication: Helicase
Unwind DNA
37
New cards
Replication: Single-Stranded DNA-Binding Proteins
Bind unraveled DNA to prevent reassociation and degradation by nucleases
38
New cards
Replication: DNA Topoisomerase
Negatively supercoil DNA to relieve positive supercoiling from DNA unwinding

Cut and reseal strands
39
New cards
Semiconservative Replication
1 retained parent strand + 1 new daughter strand makes new DNA molecule
40
New cards
Replication: DNA Polymerase
Read DNA template (parent strand) 3’ to 5’

Synthesize complementary daughter strand 5’ to 3’
41
New cards
Replication: Leading Strand
Continuous replication in replication fork direction

3’ to 5’ parent strand
42
New cards
Replication: Lagging Strand
Broken replication in opposite direction of replication fork

5’ to 3’ parent strand

Synthesize Okazaki fragments

More prone to mutations (more replication starts/stops and more primers)
43
New cards
Replication: Primase
Add RNA primer (5’ to 3’) for DNA polymerase

1 on leading, multiple on lagging
44
New cards
Replication: Prokaryote DNA Polymerase Synthesize Strands
DNA polymerase III
45
New cards
Replication: Eukaryote DNA Polymerase Synthesize Strands
DNA polymerases α, δ, ε
46
New cards
Replication: Incoming Nucleotides
5’ deoxyribonucleotide triphosphate (dATP, dCTP, dGTP, dTTP)
47
New cards
Replication: Bond Formation
Release pyrophosphate (PPi)
48
New cards
Replication: Removing RNA
Prokaryotes: DNA polymerase I

Eukaryotes: RNase H
49
New cards
Replication: Replacing RNA Primer with DNA
Prokaryotes: DNA polymerase I

Eukaryotes: DNA polymerase δ
50
New cards
Replication: DNA Ligase
Seal DNA ends
51
New cards
Replication: DNA Polymerase γ
Eukaryotes

Replicate mitochondrial DNA
52
New cards
Replication: DNA Polymerase β and ε
Eukaryotes

DNA repair
53
New cards
Replication: DNA Polymerase δ and ε
Eukaryotes

PCNA protein assists to form sliding clamp

Strengthen DNA polymerase and template strand interaction
54
New cards
Cancer Cells
From mutated genes

Excessive proliferation

Infect local (metastasis) or distant (bloodstream/lymph) tissues
55
New cards
Oncogenes
Mutated genes causing cancer (1 allele)

Code cell cycle-related proteins

Ex: Src (sarcoma)
56
New cards
Proto-Oncogenes
Pre-mutated oncogenes
57
New cards
Tumor Suppressor Genes (Antioncogenes)
Stop tumour progression

Code proteins inhibiting cell cycle or for DNA repair

Mutation loses tumor suppression activity (2 alleles)

Ex: p53, Rb (retinoblastoma)
58
New cards
Proofreading
In S phase

DNA polymerase detect instability from mismatched H bonds between incorrect paired bases

Excise and replace base
59
New cards
Proofreading: Differentiating Parent and Daughter Strands
Methylation level

Template strand older = more methylated
60
New cards
Mismatch Repair
In G2 phase

Enzymes detect and remove replication errors

Eukaryote Genes: MSH2 and MLH1

Prokaryote Genes: MutS and MutL
61
New cards
Nucleotide Excision Repair (NER) Mechanism
In G1 and G2 phases

Remove lesions distorting DNA helix

Ex: Thymine dimers caused by UV light
62
New cards
NER 1: Scan DNA
Identify bulges in strand
63
New cards
NER 2: Excision Endonuclease
Cut phosphodiester backbone to remove thymine dimer
64
New cards
NER 3: DNA Polymerase and DNA Ligase
Fill gap and seal strand
65
New cards
Base Excision Repair
In G1 and G2 phases

Remove lesions not distorting DNA helix

Ex: Uracil from cytosine deamination caused by thermal energy absorption
66
New cards
Base Excision Repair 1: Glycosylase Enzyme
Recognize and remove uracil

Leave apurinic/apyrimidinic (AP) or abasic site
67
New cards
Base Excision Repair 2: AP Endonuclease
Recognize and remove damaged sequence in AP site
68
New cards
Base Excision Repair 3: DNA Polymerase and DNA Ligase
Fill gap and seal strand
69
New cards
Recombinant DNA
DNA fragments from different sources

Gene cloning and polymerase chain reaction (PCR)
70
New cards
DNA Cloning
Produce large amounts of desired DNA sequence
71
New cards
DNA Cloning 1: Recombinant Vector
Restriction enzyme cleave vector plasmid (bacterial/viral) and DNA

Ligate DNA to vector plasmid

Transfer to host bacterium
72
New cards
DNA Cloning 2: Bacteria Colonies
Multiply DNA in vector
73
New cards
DNA Cloning 3: Isolate Colony
Include antibiotic resistance gene in recombinant vector

Grow many colonies
74
New cards
DNA Cloning 4: Express Gene
Generate large amounts of recombinant protein
75
New cards
DNA Cloning 4: Lyse Bacteria
Isolate and replicate recombinant vector

Restriction enzyme processing release cloned DNA
76
New cards
Restriction Enzymes/Endonucleases
Recognize and cleave at palindromic sequences

Isolated from source bacteria
77
New cards
Restriction Enzymes: Sticky Ends
Offset cuts

Facilitate recombination of restriction fragment with vector DNA
78
New cards
DNA Libraries
Produced from cloning

Known DNA sequence collection

Genomic or cDNA
79
New cards
Genomic Libraries
Large DNA fragments

Coding (exon) and noncoding (intron) regions

CANNOT make recombinant proteins or for gene therapy
80
New cards
cDNA (Complementary/Expression) Libraries
Reverse-transcribing mRNA

Coding (exon) regions only

CAN make recombinant proteins or for gene therapy
81
New cards
Hybridization
Joining complementary base pair sequences

DNA-DNA or DNA-RNA recognition
82
New cards
Hybridization: PCR
Produce DNA copies by hybridization without bacteria

Amplify sequence between primers
83
New cards
PCR Materials
Template DNA

Primers

Deoxyribonucleotide triphosphates (dATP, dGTP, dCTP, dTTP)

DNA polymerase
84
New cards
PCR 1: Denature
Heat and separate double helix
85
New cards
PCR 2: Anneal
Primers complementary to sequences flanking region anneal to DNA

High GC-content for H bond stability
86
New cards
PCR 3: Extension
DNA polymerase withstanding high temps add complementary nucleotides

Cool to reanneal daughter and parent strands
87
New cards
Hybridization: Gel Electrophoresis
Separate macromolecules by size and charge
88
New cards
Agarose Gel Electrophoresis
Negative DNA migrate to anode

Longer strand = slower migration
89
New cards
Hybridization: Southern Blot
Detect DNA presence and quantity
90
New cards
Southern Blot 1: Gel Electrophoresis
Restriction enzymes cut DNA

Separate by gel electrophoesis
91
New cards
Southern Blot 2: Membrane Transfer
Separated DNA transferred from gel to membrane
92
New cards
Souther Blot 3: Probing
ssDNA probes bind to complementary sequences on membrane

Probes labeled with radioisotopes/indicators for detection
93
New cards
DNA Sequencing Materials
Template DNA

Primers

DNA polymerase

Deoxyribonucleotide triphosphates

Dideoxyribonucleotides (ddATP, ddCTP, ddGTP, ddTTP) → H at C-3’
94
New cards
DNA Sequencing 1: DNA Fragments
Produce fragments terminating in dideoxyribonucleotide (H on C-3’ prevent DNA polymerase adding nucleotides)
95
New cards
DNA Sequencing 2: Gel Electrophoresis
Separate fragments by size
96
New cards
DNA Sequencing 3: Reading
Read last base for each fragment in order to identify whole DNA sequence
97
New cards
Biotech Applications: Gene Therapy
Potential cure for inherited diseases

Transfer normal gene copy in viral vector to replace mutated/inactive
98
New cards
Transgenic Animal Models
Alter germ line from transgene (cloned gene) introduction
99
New cards
Knockout Animal Models
Delete gene
100
New cards
Transgene: Fertilized Ovum
Gene injected into nucleus

Implant into surrogate mother to produce offspring with transgene germline

Study dominant gene effects