Gene Technology Flashcards
What is a Palindromic site?
its read the same forwards and backwards
eg) CTTAAG to GAATTC
What is the role of restriction enzymes?
They cut DNA at specific Palindromic sites called restriction sites
What does using a restriction enzyme leave DNA with?
sticky ends (unpaired bases)
What does reverse transcriptase do?
mRNA ——-> cDNA
What does cDNA not have?
introns (made from mRNA post splicing)
What enzyme does DNA ——> mRNA?
RNA Polymerase
Describe Artificial Synthesis of a gene.
use a gene machine to make DNA from scratch, making oligonucleotides, join theses together to make a synthetic gene.
What is an oligonucleotide?
a sequence of around 25 nucleotides
What are the three ways we can isolate target genes?
gene machine, reverse transcriptase or restriction enzymes
What is PCR used for?
to amplify a gene
What are the 4 components of PCR?
DNA sample, free nucleotides, primers, DNA polymerase
describe the 3 steps of PCR
heat up to 95 degrees
cool to 50 degrees
heat to 70 degrees
Why do we heat the sample to 95 degrees
break the H+ bonds between so they are single stranded
Why do we cool the sample to 50 degrees
allows the primer to attach by complementary base pairing to the exposed strand - making a double strand - so DNA polymerase can bind to the double stranded section
Why do we heat the sample back up to 70 degrees
there is a higher kinetic energy increasing enzyme action
why can the DNA polymerase work at high temperatures?
extracted from thermophillic bacteria
What is a primer?
Short sequence of DNA which is complementary to the target DNA
Name three ways we can isolate a target gene
restriction enzymes, Gene machine, reverse transcriptase
What charge is DNA?
negative
Describe the process of electrophoresis
attach a fluorescent label to DNA
put labelled DNA fragments into the well at the negative end
turn on current
DNA is attracted to the positive electrode
How can we distinguish the length of DNA from electrophoresis?
smaller fragments move faster (therefore further over any given time)
How can you calibrate the scale in electrophoresis?
use pieces of DNA of a known length
How can we insert the isolated target gene into a vector?
use the same restriction enzyme so that the sticky ends are complementary
DNA ligase makes recombinant DNA
How can the vector be inserted into the plasmid?
ice cold calcium chloride and a heat shock to make a transgenic/ transformed organism
How can scientists identify transformed/transgenic organisms?
marker genes, fluorescence, antibiotic resistance
Why do we then culture the transformed/ transgenic organism (bacteria)
the organism transforms/ translates the recombinant DNA
to make desired protein eg) insulin
What is gene therapy?
altering faulty alleles that cause genetic disease
How can we silence a genetic disorder caused by a dominant allele? (eg) Huntingtons)
sufferer is heterozygous
use a vector to add a DNA fragment into dominant allele
this prevents transcription
recessive allele expressed
How can we silence a recessive allele disease (eg) cystic fibrosis)
sufferer is homozygous
use a vector to add the functional allele to DNA
dominant allele will be expressed
What is Germ line gene therapy?
Change to the alleles of Gametes (currently illegal)
What is Somatic gene therapy?
Changing the alleles of body cells so offspring don’t inherit
Identify 4 possible problems with Gene therapy
alleles inserted into wrong locus
could silence the wrong gene eg) tumour suppressor
gene could be over expressed
use of gene therapy for cosmetics
identify one use of Transformed organisms in agriculture
express a protein from bacteria which is toxic to pests
advantages of genetically modifying (transforming crops to be toxic to pests
less chemical pesticide used, more efficient, lose less energy
disadvantages of genetically modifying (transforming crops to be toxic to pests
monoculture, low genetic/bio diversity,
How can genetic modification be used in Industry?
to make enzymes eg)renin (cheese) and lipases (clothes wash)
what are the advantages of making genetically modified enzymes in industry?
reduce energy, cost, fast, cheap
How can genetic modification be used in research?
make transformed pathogens to treat disease
ads of transformed pathogens in research
treat disease, pathogens will not develop resistance, reduce suffering
disads of transformed pathogens in research
could mutate and infect humans, used as bioweapons
how can genetic modification be used in Pharming/medicine?
transform bacteria to express proteins, mammals can be transformed to produce useful products in milk
ads of genetic modification be used in Pharming/medicine
cheaper than making proteins synthetically
disads of genetic modification be used in Pharming/medicine
possible unexpected side affects eg) cancer
using animals as commodities
What is a DNA probe?
a short sequence of single stranded DNA with a label attached, that is complementary to a specific allele or gene
Describe the process of using a DNA probe
attach label to the DNA probe
If complementary DNA is present the probe will hybridise to it
rinse to remove hybridised DNA probes
View labels to establish is DNA is present
Describe the use of a DNA Microarray
label is attached to the patient DNA
can test multiple alleles/ mutations at once
many probes are attached to a tile in a grid
DNA will hybridise by complementary H+ bonding
rinse
How is Genetic Counselling used?
Healthcare professionals can advise patients about risks involving heritable disease, and risk of developing genetic diseases later in life eg) BRCA 1 gene and breast cancer
Genetic Counselling allows patients to make informed decisions about actions to take
How is Genetic Screening used?
parents see is they are carriers of recessive alleles, can also be used to diagnose and treat genetic diseases before symptoms show eg) babies and CF
How can DNA probes be used to give personalised medicine?
If a doctor is aware of genotype can give most effective drug, or drug least likely to cause side affects
What is genetic fingerprinting?
Identifying individuals by comparing differences in their Variable Number Tandem Repeats (VNTRs)
where are VNTRs located?
non - coding DNA?
Why do we use VNTRs to identify criminals in forensic investigation?
they don’t affect phenotype, so vary far more between individuals
Describe the forensic process in identifying suspects
collect samples of DNA from crime scene and suspects
amplify using PCR
run using gel electrophoresis
compare the results of suspects with the crime scene DNA
what else can genetic fingerprinting be used for?
paternity tests, establishing relatedness (used in conservation of endangered species)
