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Stage 2 Biology > DNA AND PROTEINS > Flashcards

DNA AND PROTEINS Flashcards

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1
Q

Define Phenotype

A

Physical characteristic

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2
Q

Define genotype

A

Genetic expression

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3
Q

Can alterations occur to the phenotype that don’t effect the genotype?

A

Yes

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4
Q

What is siRNA and its role?

A

Small interfering RNA prevents translation of a gene by cutting mRNA after transcription or binding to DNA and attracting other molecules to the sites of transcription and translation

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5
Q

What environmental factors can affect phenotypic expression?

A
  • Lack of oxygen
  • increased UV exposure
  • lack of iodine
  • malnutrition
  • increased light intensity
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6
Q

Describe how methylation affects the expression of a gene.

A
  • Methyl groups attach to cytosine on the promotor region of the gene.
  • They stop transcription factors from working such as RNA polymerase, therefore the desired gene is silenced as it is not expressed.
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7
Q

Describe methylation of histones.

A
  • Methyl groups attach to histones making them more ‘sticky’ (more strongly attracted to one another)
  • Transcription factors can’t bind to DNA, therefore gene is not expressed.
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8
Q

Describe Acetylation

A
  • Acetyl groups attach to histones and separate them.
  • Transcription can easily occur thus the gene is expressed.
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9
Q

Describe de-methylation

A
  • There are no methyl groups on the promotor region of the gene.
  • Transcription factors can easily bind, thus the gene is expressed.
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10
Q

Define epigentics

A
  • can be caused by environmental factors
  • can create heritable changes
  • does not involve changes to the underlying DNA sequence
  • epigenetic factors are caused by methylation
  • can cause cancer
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11
Q

Missense mutation

A

One base has changed - could influence amino acid

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12
Q

Nonsense mutation

A
  • change in nucleotide
  • changes protein
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13
Q

Insertion/deletion mutation

A

Adding/deleting a nucleotide
changes everything after

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14
Q

Causes of mutations

A
  • induced by environmental factors: ionizing radiation
  • virus
    -mutagenic chemicals
  • could occur spontaneously, generally in DNA replication
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15
Q

Difference between somatic and germ cells

A

Germ:
- effect zygote, and effects every cells DNA in the organism produced
- passed down

Somatic:
- effects an individual organism

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16
Q

Process of PCR

A
  1. DNA is extracted and placed in a test tube. DNA is heated to break hydrogen bonding and sperate into two strands (DNA denatured).
  2. DNA is cooled and primers are added complementary to the DNA sequence being copied to ensure the DNA doesn’t re-zip. Free complimentary nucleotides are added that bond to the exposed bases.
  3. DNA is heated so DNA polymerase and continue DNA replication, creating two new complementary DNA strands by connecting the sugar phosphate backbone.
  4. DNA is cooled so hydrogen bonds can rejoin, and process is repeated.
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17
Q

What is in the centromere of our chromosomes?

A
  • variable number tandem repeats (VNTR’s)
  • non coding pieces of DNA
  • This makes each human an individual, the parts of DNA that separate us.
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18
Q

How are VNTR’s analysed?

A
  • Amplified through PCR
  • Analyzed through gel electrophoresis, creating bands of DNA in the gel.
    Individuals have unique banding patterns
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19
Q

Define the process of electrophoresis

A
  1. DNA samples are separated into ‘wells’ at the negative end of the plate.
  2. DNA is negatively charged, this travels to the positive end.
  3. Dependent on the size of the DNA fragment (number of VNTR’s you have), shorter strands will go further up the plate towards the positive end whereas larger pieces will get stuck within the gel.
20
Q

Define Electropherograms

A

The same electrophoresis only the end nucleotide of each piece of DNA is labelled with a fluorescent dye.
1. DNA passes through a capillary tube containing the gel and a laser readers the dyed fragments as they pass.

21
Q

STR

A
  • Short tandem repeats (STR)
  • Replacing VNTR’s
  • regions scattered throughout the genome which are made of repeated sequences between 2-8 nucleotide base sequences.
  • Each individual has two sets of chromosomes, so you have two STR numbers e.g., (11, 12).
22
Q

DNA profiling

A
  • electropherogram is used to make a DNA profile of an individual.
  • Increasing the number of loci where STR’s are located, you can see more of the uniqueness of an individual
  • Information can be represented on a graph to identify the chromosome locus and the number of allele repeats.
23
Q

Ethical considerations of DNA Profiling

A
  • could purposely create DNA sequences that are harmful/dangerous
  • concerns that an individuals’ genetic blueprint can be recorded/altered.
  • life insurance companies could check a persons genome for potential future illnesses and refuse insurance or charge them more.
24
Q

Define Recombinant DNA

A
  • Talking DNA from one species and putting it in another.
25
Q

Define restriction enzymes

A
  • proteins that come from bacteria and cut DNA at highly specific regions/sequences.
26
Q

Define a restriction site

A
  • the sequence of bases recognized by a restriction enzyme.
  • some restriction enzymes cut straight across the double helix causing blunt ends, others create sticky ends.
  • sticky ends leave a few nucleotides so when the new gene is inserted it can bind through hydrogen bonding.
27
Q

What are two ways of isolating genes?

A
  • Probing DNA
  • Antibodies
28
Q

What is a probe?

A
  • A short piece of single stranded DNA or RNA
29
Q

Probing DNA

A
  • The sequence of nucleotides that the probe is looking for must be known and can be identified using the amino acid sequence of a protein.
  • The probe has the complimentary strand of the nucleotides and is labeled radioactively.
  • Radioactively labeled probe binds to the desired gene that is being looked for.
30
Q

Define the antibody method of isolating a gene.

A

STEP 1: CREATE THE ANTIBODY
- The protein that is coded for by the gene of interest is injected into an animal.
- Animal makes antibodies because it is a foreign entity (protein).
- Antibodies are harvested from blood and labeled either radioactively or chemically (using a fluorescent marker).
STEP 2:
- DNA from cell is cut up into fragments and incorporated into bacteria cells.
- Some bacteria will take this DNA and express the proteins related to the genes.
- The labelled antibody is applied to the bacteria
- The bacteria that have taken the gene will be identified by the label, the ones that haven’t taken the other genes will not be fluorescent.
- The labeled bacteria can be harvested and cloned.

31
Q

Why do we clone genes?

A
  • Used to transfer a gene from one species to another.
32
Q

The process of cloning genes

A
  1. Restriction enzymes cut plasmids (in bacterial DNA), leaving the sticky ends.
  2. The gene we want is inserted by mixing it in and the complementary sticky ends adhere to the plasmid. - DNA ligase rejoins the plasmid into rings.
  3. New plasmids are introduced to a bacterial colony which might be taken up.
  4. Reproduced the desired gene.
  5. Insert into the cells that we want the gene to go into.
33
Q

Transgenics (how to get the gene of interest into cells)

A
  • Transfection vectors (viruses, liposomes)
  • electroporation
  • gene gun
  • microinjection
34
Q

Define Gene Gun

A
  1. Coat inert particles (like gold) with the gene of interest.
  2. This fired under high pressure at the cells we want to put the gene into.
  3. Some particles will enter cells and some cells will incorporate the new gene into the genome.
  4. These cells then express the desired protein.
35
Q

Define Agrobacterium

A
  • causes tumors in plants
    1. The Ti plasmid is removed from bacteria and the desired gene we want added.
    2. The plasmid is returned to the bacteria and the bacteria is used to infect a plant.
    3. Bacteria inserts the Ti Plasmid (containing our gene) into cells, which divide rapidly to produce a tumor.
    4. The tumor is then harvested, cut up an used to create plants with our gene as a part of the genome.
36
Q

Define Electroporation

A
  • A short electric shock is applied to the plasma membrane of a cell, creating small pours for the particles containing the desired gene to pass through the membrane into the cell.
37
Q

Vectors: Gene therapy

A

Virus:
1. Modify DNA the virus carries to be the one with the gene we want expressed and remove part that tells the cell to keep replicating.
2. Virus invades host cell and inserts DNA in the cell.

38
Q

Microinjection

A
  1. A very fine micropipette is inserted through the plasma membrane and the gene of interest is inserted into the nucleus of the ovum. Therefore, organism has the gene of interest.
39
Q

How does CRISPR work in bacteria?

A
  • bacteria store some viral DNA in Clustered Regularly Interspaced Short Palindromic Repeats.
  • if the same virus attacks at a later date, the bacteria recognizes this and creates guide RNA complimentary to the viral DNA copy.
  • RNA is loaded into the Cas-9 enzyme which looks out for its complimentary pair, and when viral DNA does invade, it will cut it.
40
Q

How does CRISPR work?

A
  1. Guide RNA is loaded into the Cas9 enzyme.
  2. Cas9 travels along the genome and attaches itself to the segment of DNA complimentary to the guide RNA.
  3. The restriction enzyme cuts the DNA to deactivate the gene.
  4. The new gene is mixed into solution and inserts itself when DNA repairs itself with the new gene. Ligase seals DNA back up.
41
Q

Designer proteins

A
  • Look at the protein we want to make.
  • Determine amino acid sequence, therefore DNA sequence can be determined
42
Q

What can designer proteins be used for?

A
  • Vaccines - that bind to viruses and bacteria to prevent them from working.
  • protein spheres - similar to liposomes that an deliver DNA payloads to cells.
  • Channel proteins that regulate the movement of substances through the cell membrane.
  • proteins that could glow when they detect specific molecules.
43
Q

Genetic engineering for use of vaccines

A
  • The protein from the viruses coat can be produced through recombinant technology and injected into an animal as an antigen which stimulates immune response and the production of antibodies. These antibodies can be used to treat people with that particular virus.
  • The replication portion of the viruses genome can be removed from the viruses genome and inject replication-deficient virus to stimulate an immune response.
44
Q

Explain how enzymes work as catalysts

A
  • enzymes have an active site which the substrate is complimentary to.
  • the substrate binds with to the enzyme, and the active site changes shape slightly to place stress on bonds holding the substrate.
  • The stress becomes too much forcing the substrate out of the active site where the bonds have broken but new ones form, creating a new product.
  • Enzymes reduce activation energy a the correct orientation is provided for the reaction to happen.
45
Q

Process of gel electrophoresis

A
  1. Restriction enzyme cuts DNA
  2. DNA is placed in agar
  3. Electricity is passed through
  4. Smaller fragments travel closer to the positive electrode
46
Q

Describe the process of DNA replication

A
  1. Helicase separates the DNA molecule into two strands by breaking the hydrogen bonds.
  2. Free complimentary nucleotides attach to either strand of the exposed bases.
  3. DNA polymerase connects the sugar-phosphate backbone to form two DNA molecules.
47
Q

Process for microinjection in an embryo

A
  1. Locate gene of interest
  2. Isolate gene
  3. Use PCR to copy gene
  4. Isolate zygote
  5. Transfer copies of the gene into a fertilized egg
  6. Express the gene of interest.

Stage 2 Biology (3 decks)

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