Week 1 Flashcards
1
Q
What is Heredity?
A
Process in which traits pass from one generation to the next one without being altered.
2
Q
Why is heredity important?
A
DNA keeps the fidelity of those traits.
3
Q
What happens when DNA doesn’t keep the fidelity of various traits?
A
when this is not happening changes in DNA cause modifications.
4
Q
What are the biological functions of proteins?
A
Responsible for most biological functions like Structure, movement etc.
5
Q
What is the purpose of DNA?
A
Stores heritable information.
Set of instructions to build and maintain an organism is stored in the DNA.
Provides continuity between generations.
Is the substrate for evolution (Natural selection is going to try and preserve those alleles.
6
Q
Where is DNA found in eukaryotic cells?
A
Nucleus
Mitochondria
Chloroplasts
7
Q
Where is DNA found in prokaryotic cells?
A
Circular chromosome in the nucleoid
Plasmids
8
Q
What are the building blocks of DNA?
A
Nucleotides
Nitrogenous base
Pentose sugar
Phosphate group
9
Q
What is created Without the phosporus group?
A
nucleosides
10
Q
What are the four bases?
A
guanine, adenine, Cytosine, thymine, (uracil)
11
Q
What are the 4 nucleosides?
A
guanosine, Adenosine, Cytidine, thymidine, (uridine)
12
Q
What are the 4 nucleotides?
A
guanylate, Adenylate, Cytidylate, thymidylate, (uridylate)
13
Q
How are nucleotides linked?
A
by phosphodiester bonds
14
Q
Pyrimidines contain what?
A
Cytosine
Thymine (T in DNA)
Uracil (U in RNA)
15
Q
What do Purines contain?
A
Adenine
Guanine
16
Q
What does the DNA strand contain?
A
All have a sugar-phosphate backbone.
Has directionality 5’ to 3’
17
Q
How was the structure of the double helix discovered?
A
Structure of DNA was solved by Watson and Crick using X-ray data collected by Rosalind Franklin
18
Q
What are the features of the double helix?
A
Relatively hydrophobic nitrogenous bases in the molecule’s interior
Negatively charged phosphate groups in the exterior
Sugar-phosphate backbone
Weak hydrogen bonds hold the two strands together
19
Q
What are the base pairings?
A
“Chargaff’s ratios”: Ratio of A : T and G : C bases always the same
This is because A always pairs with T, and C with G
20
Q
What are bacterial chromosomes?
A
Usually a single circular DNA molecule
DNA packed with a small amount of proteins
21
Q
What are Eukaryotic chromosomes?
A
The genome is found in multiple chromosomes
Each chromosome contains is a linear DNA double helix
DNA packed with a large amount of proteins (this is called chromatin)
22
Q
Why is packing necessary?
A
DNA is very long.
Long DNA molecules in chromosomes need to be packed so they fit into the cell.
Average length of DNA in human chromosomes: 1.5 x 108 base pairs
Length of 1 DNA molecule = ~ about 4 cm
There are 46 DNA molecules in a human cell
4cm * 46 = Total length of DNA 184 cm
23
Q
How is DNA packed?
A
Packed by histone proteins. Histones leave DNA briefly, during DNA replication and transcription
24
Q
Chromosomes are NOT _______ organised in the nucleus.
A
Randomly.
25
What is active chromatin?
Euchromatin: loose chromatin structure, active for transcription (DNA transcribed to RNA).
26
What is inactive chromatin?
Heterochromatin: condensed chromatin structure, inactive for transcription.
27
What is a gene?
Sequence of nucleotides that encodes the synthesis of a gene product, either RNA or protein
The order of the nucleotides determines the order of monomers in a polypeptide or nucleic acid molecule
28
What does the promoter on the gene do?
(controls expression of
| the gene) can be switched on or off.
29
What do genes have at the 5' end?
Genes have a start codon
| = ATG nucleotide sequence
30
What do genes have at the 3' end?
Genes have a stop codon
| (several different sequences
31
What is a genome?
The complete set of genetic material present in a cell or organism
32
What is genomics?
Study of the whole genomes.
33
What does genomics contain?
Sequencing of genomes
Study of evolution of genomes
Study of regulation of whole genomes
Behavioural genomics
34
What is transcriptomics?
Study of all the mRNAs present in an organism or cell.
35
What is Proteomics?
Study of all the proteins present in an organism or cell.
36
What is Metabolomics?
Study of all the metabolites in an organism or cell.
37
What is Epigenomics?
Study of all the epigenetic state of the whole genome.
38
What is a genetic locus?
A particular location on the genome or on a chromosome.
Most often refers to a gene
But it can also refer to just one nucleotide in the genome (e.g. that has a mutation)
It can be used to refer to a whole gene
It can be used to refer to other bits of DNA that are not genes
39
What is semi-conservative replication?
Each strand serves as the template for replication of a new strand of DNA
40
What is the origin of replication?
DNA replication starts at a specific location in the genome
41
How many origins are in bacterial chromosomes?
Only 1
42
How many origins are in eukaryotic chromosomes?
May have hundreds or even thousands of origins
43
What is the very active area where DNA replication takes place known as?
Replication fork or bubble
44
How do replication forks move?
In opposite directions.
45
What is required for DNA replication?
Multiple enzymes
46
What occurs due to Topoisomerase?
unwinds DNA, helix to ladder
47
What occurs due to Helicases?
unzips DNA, double-stranded DNA (dsDNA) to single-stranded DNA (ssDNA)
48
What do Single-stranded binding proteins do?
stabilizes ssDNA
49
What do primase do?
RNA polymerase, sets RNA primer to start new strand.
50
What does DNA polymerases do?
adds new nucleotide based on complementarity with DNA template.
51
What do DNA ligase do?
links the nucleotides in the new strand
52
What can DNA polymerases not do?
Cannot start a DNA chain, only extend it.
53
What does DNA pol III add?
nucleotides to the RNA primer
54
Which direction does DNA polymerases add nucleotides in?
5’ to 3’ direction
55
Why do nucleotides need to be added in the 5' to 3' direction?
Otherwise, later proofreading wouldn’t have access to the energy of the 2 phosphates used in the ligation.
56
What happens when DNA polymerase edit the wrong base?
causes mutations
57
During DNA duplication, ________ __________ can check each base that they add.
DNA polymerase
58
During DNA proofreading what does DNA polymerases compare?
compare the 2 paired bases, and if they don't follow the A-T or C-G pairing, then they edit one to make them to match.
59
Outline the process of DNA proofreading.
1) Polymerase adds an incorrect nucleotide to new DNA strand.
2) Polymerase detects that bases are misplaced.
3) Polymerase uses 3' to 5' exonuclease activity to remove incorrect nucleotides.
60
What role does the Topoismerase have in the replication fork?
Breaks, rotates, and then rejoinds the DNA
61
What role does the Helicase have in the replication fork?
Unwinds/separates parental strands
62
What role does the primase have in the replication fork?
makes RNA primers, using parental DNA as template
63
Due to the limitation of only building strands from 5' to 3', what will happen to some strands?
some strands will be synthesised very fast known as leading strands and strands known as lagging strands.
64
How long are the Okazaki fragments in E.coli and eukaryotes?
1,000-2,000 bp long in E.coli
| 100-200 bp long in eukaryotes
65
What does DNA ligase do in antiparallel elongation?
joins the sugar-phosphate backbones of the Okazaki fragments
66
What does DNA pol I do in antiparallel elongation?
replaces the RNA nucleotides of the adjacent primer with DNA nucleotides
67
During the Replication of Chromosome Ends, what happens to the linear chromosomes?
get shorter after each round of replication
68
Where is replication not fully efficient?
In eukaryotes.
69
What are Telomeres?
the ends of eukaryote chromosomes
Consist of many short repeats
E.g. TTAGGG in humans
70
How are Telomeres added?
These repeats are added on by telomerase
71
What is genetic engineering?
Set of technologies used to change the genetic makeup of cells, including the transfer of genes within and across species boundaries
72
What research has gone into genetic engineering?
loss of function experiments (gene knockout), gene gain of function, Green Fluorescent Protein (from jellyfish).
- Medicine- mass production of insulin inserted in bacteria.
- Industry- making biofuels, cleaning up oil spills, carbon and other toxic waste.
- Agriculture- pest resistant crops and golden rice (vitamin A).
- Cloning.
73
What are the typical gene cloning strategies?
1) Isolate gene/promoter from organism.
2) Manipulate it in vitro.
3) Analyse its function.
74
How is a gene isolated?
Through PCR.
75
Why is PCR useful?
- We need multiple copies of a gene to be able to study it
- PCR is a simple technique to make many copies of a specific piece of DNA
- It can be used to isolate a gene from the genome of an organism
76
Who invented PCR?
By Kary Mullis in 1983
77
What are the ingredients needed for PCR?
Template DNA
Primers
Nucleotides
DNA polymerase
78
How are Primers used in PCR?
Primers flanking ends of DNA section that will be amplified and isolated
79
How are free nucleotides used in PCR?
A, G, C, T to build DNA
80
During PCR DNA template needs to be _______. e.g. DNA isolated from ______.
blank 1- amplified
| blank 2- animal
81
What are the three stages of PCR?
Denaturalisation.
Annealing.
Extension.
82
What happens during denaturalisation?
Heat DNA to 95 °C for 1 minute to unzip the double strands
83
What happens during Annealing?
cool to ~60 °C to allow short ‘primers’ to bind to DNA
84
What happens during Extension?
Heat to 72 °C, the optimum temperature for a DNA polymerase
85
How can the PCR produced template be used?
Using as a DNA barcode species.
86
What is present within the third domain in the universal tree of life?
In the third domain (Eukarya) multicellular life is largely restricted to three recent branches (fungi, plants, animals)
87
2 of the 3 domains in the universal tree of life are populated by what?
domains populated exclusively by prokaryotes
88
How is the gene cloned?
Mannipulate it in vitro.
89
Summarise gene cloning.
1) Human genome.
2) Gene of interest isolated from the genome- PCR or restriction enzyme.
3) Gene inserted into plasmid.
4) Plasmid inserted into bacteria population generating "clones" of the gene.
90
What is E.coli an important tool for?
Gene cloning
91
What are plasmids?
are small circular pieces of DNA found in bacteria
92
Gene Cloning-
| We can _____ genes in plasmids, and then transform plasmids in ______ .
Insert
| Plasmids
93
What happens to the transformed bacteria in gene cloning?
transformed bacteria divide making copies of the plasmid inside them.
94
When is gene cloning used?
Used before PCR, but still a main lab technique in genetics
95
Plasmids are = to ____.
Cloning vectors
96
We will ______ our gene of interest in the plasmid using _____ _______.
blank 1- insert
| blank 2- restriction enzymes
97
What do we need to check during gene cloning?
We need to check that gene was inserted in the plasmid
98
What happens once the we check that gene was inserted in the plasmid?
Then we will transform the plasmid (introduced it within a bacterial cell)
We need to check that plasmis was introduced in the bacteria
99
What are restriction enzymes?
type of endonucleases
100
What do restriction enzymes recognise and what are there functions?
recognize specific short sequences of DNA and
| Then they cleave the DNA at or near that site
101
How long are recognition sites?
are usually 4-8 bp long and palindromic – same on both strands
102
What are the roles of restriction enzymes in nature?
In nature, these enzymes protect bacterial cells against invasion by viral DNA
103
How is bacterial DNA protected?
is protected by addition of methyl groups
104
How is each enzyme named?
Each enzyme is named after its organism of origin
105
What do all these restriction enzymes do?
All these enzymes recognize and cut specific but different DNA sequences
Researchers select and order right enzyme for the job
106
Summarise what restriction enzymes do in bacterial plasmids.
1) Restriction enzyme digests(cuts) the sugar-phosphate backbones at each arrow.
2) Base pairing of sticky ends produces various combinations.
3) Fragment from different DNA molecule cut by the same restriction enzyme
4) DNA ligase seals the strands. Recombinant DNA molecule
5) Recombinant plasmid
107
What is transformation?
introducing the plasmid in the bacteria
108
What are the two types of transformation?
Chemical and Electroporation
109
What is chemical transformation?
Incubation
heat shock
return to ice
All opens up the membrane to let plasmid in.
110
What is electroporation?
Electric shock- return.
111
What are the two checks for gene cloning?
- We need to check that plasmid was transformed in the bacteria
- We need to check that our gene was inserted in the plasmid
112
How can we check that the plasmid was transformed in the bacteria?
- Plasmid contains antibiotic resistance gene
- Bacteria are grown in a medium with antibiotic
- Only bacteria with the plasmid can grow, the bacteria with no plasmid die because of the antibiotic
113
How can we check that our gene was inserted in the plasmid?
- Plasmid cloning site is within a protein coding gene (lacZ)
- This gene produces pigmented colonies (e.g. blue)
- If no gene of interest is inserted, color gene does work, then colonies have colour (e,g, are blue).
- If our gene of interest is inserted, the colour gene is broken, then bacteria are white
114
What are the features of a typical gene cloning vector?
- Antibiotic resistance gene with bacterial promoter
e. g. b-lactamase
- Multiple cloning site” in colour gene
- Origin of replication
115
How do you cut and paste with restriction enzymes?
For example- using GFP to study location of a protein in a cell
A gene encoding a protein under investigation
(isolated with PCR)
-In the lab, the GFP gene is linked to the gene encoding the protein
(using restriction enzymes)
-
116
How is bacteria cloned?
Gene inserted into plasmid
Plasmid put into bacterial cell.
Host cell grown in culture to form a clone of cells containing the cloned gene of interest.
117
How do we analyse the function, and separate and visualize DNA ?
Gel electrophoresis
118
Why are Nucleic acid molecules attracted to a positive charge?
Nucleic acids (DNA and RNA) are acid (they have a negative charge)
119
What is Agarose gel?
Agarose is a polysaccharide from red algae
Liquid at high temperature, solid at room temperature leads to casting of the gel.
120
outline a few steps of gel electrophoresis.
- Agarose in container is cooled down.
- Mix DNA with blue dye makes it visible.
- Cathode and anode used to make electrical current and electrical field to allow DNA to move.
121
How is DNA separated and visualized through gel electrophoresis?
1) DNA for analysis is pipetted into small holes in the gel
(called ‘wells’)
2) A gel of an aequeous polymer
(usually agarose)
3)DNA migrates through the gel in response to an electrical field
4)The pores in the gel act as a sieve, so smaller DNA molecules travel faster and further
122
What does the matrix in the agarose make easier?
For smaller DNA fragments to travel faster.
123
How is DNA visible during gel electrophoresis?
the gel is stained with a DNA intercalating dye
Ethidium bromide is a large flat molecule
It can bind to DNA
It is fluorescent (visible under UV light)
124
What are the three models of DNA replication?
Conservative model
semi-conservative model
Dispersive model
125
what is the conservative model?
Both strands of parental duplex would remain intact and new DNA copies would consist of all new molecules, Daughter strands contain all new molecules.
126
what is the semi- conservative model?
1 parental duplex remains intact in daughter strands, a new complementary strand is built for each parental strand consisting of new molecules. Daughter strand- 1 parental strand and 1 newly synthesized strand
127
What is the dispersive model?
DNA consists of mixtures of parental and newly synthesised strands new DNA would be dispersed throughout both daughter molecules after replication.
128
Replication requires three things-
1) Something to copy.
2) Something to do the copying.
3) Building blocks to make the copy.
129
What are the Seven characteristics shared by living systems?
```
Cellular organization
Ordered complexity
Sensitivity
Growth, development and reproduction
Energy utilization
Homeostasis
Evolutionary adaptation
```
130
What is the cellular level?
At the cellular level we have atoms and molecules assemnled into organelles within cells which are the basic units of life.
131
The organization of the biological world is _______= each level builds on levels below it.
hierarchical
