manipulating genomes and genetic engineering

Question 1

what is a DNA probe

Answer 1

a small complimentary section of code of a gene you are looking for. contains fluorescent marker or radioactive dye.

Question 2

what are primers

Answer 2

short pieces of single stranded DNA that are complimentary to the bases at the start of the DNA fragment you want.

Question 3

outline the 3 stages of PCR

Answer 3

1. heat to 95 degrees, breaks hydrogen bonds between 2 strands of DNA
2. cool to 55 degrees, allows primers to anneal
3. heat back to 72 degrees, free complimentary nucleotides are joined by taq polymerase which forms sugar phosphate backbone

Question 4

what are the 2 methods scientists can use to isolate a gene

Answer 4

restriction endonucleases, - restriction enzymes that cut at specific sites isolating the gene and leaving sticky ends. different restriction enzymes cut at different specific sites.

reverse transcriptase which converts MRNA of complimentary gene to cDNA

Question 5

outline the steps of setting up gel electrophoresis

Answer 5

gel tray placed into gel box. wells are at negative (cathode) end. buffer solution is added to reservoirs to ensure solution carries current + maintain pH.
DNA sample is loaded into each well using a micropipette and loading dye is added to ensure the sample sinks making it easier to see.
place lid on box and connect to power supply, turn on and leave electrical current to pass through gel

Question 6

what is the gel called in gel electrophoresis

Answer 6

agarose gel

Question 7

which way will DNA fragments move in gel electrophoresis and why

Answer 7

will move from the cathode to the anode as DNA fragments are negatively charged (phosphate group) and so travel to positive end.

Question 8

how are DNA fragments separated in gel electrophoresis

Answer 8

by size, smaller fragments travel further.

Question 9

what is done with proteins before gel electrophoresis is used and why

Answer 9

denatured with a chemical so that they all have the same charge

Question 10

what does southern blotting involve

Answer 10

nylon sheet is placed on top of gel trat, DNA fragments are soaked onto nylon sheet

Question 11

what is hybridisation

Answer 11

fluorescent dye is added to fragments and they are viewed under UV light
or
radioactively labelled dye is added to fragments and viewed under X-ray.

Question 12

what is a genome

Answer 12

all the genetic material in an organism

Question 13

which is smaller microsatellites or minisatellites

Answer 13

microsatellites

Question 14

what is unique in each persons genome that allows DNA profiling to occur

Answer 14

satellite pattern - these are non coding sequences that repeat at a specific loci, the number of repeats is unique to each individual so each person has a different satellite pattern.

Question 15

what are the different uses of DNA profiling

Answer 15

used in forensic analysis to compare crime scene evidence to potential suspects

used in paternal cases

used in medical diagnosis - analyses the risk of genetic disorders
eg. compares DNA profile of embryo to parents DNA profile, if the profiles match the embryo has inherited the disorder. - used to screen for cystic fibrosis.

Question 16

what are transformed organisms

Answer 16

organisms that have recombinant DNA - DNA formed by joining together DNA from different sources.

Question 17

what is a transgenic organism

Answer 17

an organism that has been genetically engineered to include a gene from a different species

Question 18

what is a plasmid

Answer 18

small circular molecule of DNA in bacteria

Question 19

what is a bacteriophage

Answer 19

virus that can infect bacteria

Question 20

outline the genetic engineering process

Answer 20

• isolate desired gene/DNA fragment using restriction enzymes
• vector DNA (plasmid) is cut upon using the same restriction enzymes (so sticky ends are complimentary)
  -vector DNA and DNA fragment is mixed together with DNA ligase - forms sugar phosphate backbone
  -the vector is used to transfer the gene into bacterial cells, host cells are persuaded to take in the plasmid by electroporation/heat shock/electrofusion

Question 21

how does a bacteriophage vector transfer the gene to the host

Answer 21

infects the host bacterium by injecting its DNA into it, the phage DNA then integrates into the bacterial DNA

Question 22

what happens during heat shock

Answer 22

plasmid and bacteria are placed into solution containing Ca2+ ions, temperature is increased. this increases permeability of bacterial membrane + kinetic energy

Question 23

in genetic engineering of plants, what is the name of the bacterium in which the plasmid is taken from

Answer 23

Argobacterium tumefaciens

Question 24

list 3 advantages of genetically modified crops

Answer 24

1. reduces amount of pesticides farmers have to use which can harm environment (if insect resistent modified)
2. GM plants can be designed to be more nutritious
3. increases the crop yield

Question 25

list 3 disadvantages of genetically modified crops

Answer 25

1. may encourage monoculture - decreases biodiversity
2. if all genetically identical whole crop is susceptible to disease
3. could interbreed with wild plants creating ‘super weeds’

Question 26

what is name of the term given to producing drugs from animals

Answer 26

pharming

Question 27

outline how the protein antithrombin can be produced by goats

Answer 27

1. DNA fragments that code for production of protein in mammary gland are extracted
2. dna fragments are injected into a goat embryo
3. embry implanted into surrogate mother
   4.if offspring produces the protein, selective breeding is sused to produce a herd of goats that produce the protein in their milk. which can be extracted

Question 28

what are 2 benefits of pharming

Answer 28

1. drugs can be made in large quantities
2. this makes them more available to people

Question 29

what are 2 disadvantages of pharming

Answer 29

concern manipulating animal genes could have. harmful side effects for animal

enforces idea that animals are merely ‘assets’ we can use.

Question 30

how have pathogens been used in research

Answer 30

tumour cells have receptors on membranes for poliovirus meaning the virus recognises and attacks tumour cells.

scientists have. manipulated the poliovirus to inactivate the genes that cause the disease, so can use poliovirus to attack and kill cancer cells without infecting the person

Question 31

what is an advantage of using pathogens in research

Answer 31

previously untreatable diseases can now be treated, reduces the suffering. they cause

Question 32

what is a disadvantage of using pathogens in research

Answer 32

some people are worried it could cause an outbreak

The GM version could revert back to origional form and cause outbreak

in wrong hands could be used maliciously to create agents for biowarfare.

Question 33

what are the 2 types of gene therapy

Answer 33

somatic
germ line

Question 34

how would a genetic disease caused by 2 recessive alleles be treated

Answer 34

a working dominant allele could be added. eg by using vectors

Question 35

how would a genetic disease caused by a dominant allele be treated

Answer 35

by silencing it - sticking a bit of DNA in middle of allele so it doesn’t work anymore.

Question 36

why is a virus the most common vector used in genetic engineering

Answer 36

it has the mechanism to recognise cells and deliver genetic material into them.

Question 37

what is somatic therapy

Answer 37

inserting functional alleles into body cells, can be:
ex-vivo: new gene is inserted via a virus vector into cells outside the body. eg. bone marrow cells are extracted and exposed to virus which inserts gene into cells. these cells are grown in a lab and returned via injection

in vivo: new gene is inserted via a vector into cells inside the body.

Question 38

is somatic therapy long term, can offspring inherit the genetic disorder

Answer 38

somatic therapy is not long term as only alters body cells which die and become replaced, therefore patients need to return for multiple treatments.

somatic therapy doesn’t affect the individual sex cells so any offspring can still inherit the disorder

Question 39

what is germ line therapy

Answer 39

inserting functional alleles into zygotes or gametes - every offspring produced will be alered by the gene therapy.

Question 40

what is DNA sequencing

Answer 40

determining the sequence of nucleotides in a piece of DNA

Question 41

what are terminator bases

Answer 41

bases that have a hydrogen atom instead of OH group on 3’ end. this means they cannot form phosphodiester bonds with the next nucleotide .

Question 42

outline the steps of pyrosequencing

Answer 42

1. section of DNA is cut into fragments, a strand from each fragment is attached to a small bead
2. PCR amplifies the DNA fragments on each bead
3. each bead is placed into a separate well
4. free nucleotides are added
5. wells contain specific enzymes which cause light to be emitted when a nucleotide is added
6. computers analyse the occurrence and intensity of light and process to interpret DNA sequence