Genome Projects and Gene Technologies Flashcards
define genome
complete set of genetic material that organism has
define proteome
complete set of proteins that organism can make
human genome project
- sequenced whole genome
- only sequence short fragments at once
- split genome into small sections - sequence - put back in order
how are sequencing methods being constantly updated
- chain termination method
to
- pyrosequencing
- automated
- faster cheaper
predicting AA sequences in simple organisms
- few regulatory genes
- little non coding DNA
predicting AA sequences in complex organisms
- non-coding dna e.g. VNTRs
- regulatory gene turns on/off other genes
- lots of non-coding dna
- hard to determine proteins from dna
uses of genome projects
- understand evolutionary relationships
- medicine - understand antigens to develop new vaccines
define gene therapy
changing faulty alleles that cause genetic disease
gene therapy for disease caused by dominant allele
- sufferes = hetero
- alreadu have functional allele
- silence dominant allele
- use vector to add dna fragment into dominant allele
- dominant allele not transcribed
- recessive allele expressed
gene therapy for disease caused by recessive allele
- sufferers = homo
- use vector to add functional allele to DNA
- dominant allele expressed
2 types of gene therapy
- germ line
- somatic
germ line gene therapy
- changing alleles of gametes
- all future offspring inherit
- illegal
somatic gene therapy
changing allele of body cells
- offspring dont inherit changes
problems with gene therapy
- alleles inserted into wrong locus
- could silence wrong gene by mistake
- could cause cancer
- gene over expressed
- non-medical uses
3 ways to isolate target genes
- restriction enzymes
- reverse transcriptase
- gene machine
restriction enzymes
- RE cut dna at specific palindromic sites = restriction sites
- if restriction site either end, RE used to cut it out
- leaves dna with sticky ends (unpaired bases)
reverse transcriptase
- cells have 2 copies of each gene - hard to access
- enzyme that does transcription backwards
DNA + RNA polym –> mRNA
mRNA + reverse transcriptase —> cDNA
gene machine
- join 25 nucleotides together at once
- forms oligonucleotides
- join oligonucleotides = synthetic gene
- design own genes/ proteins
what is PCR used for?
- amplify DNA (make millions of copies)
known as IN VITRO dna amplification
PCR ingredients
- DNA sample
- free DNA nucleotides
- primers
- DNA polymerase (taq polymerase)
what are primers?
short sequences of DNA that are comp to start of dna sample
- used to select which part of dna copied
PCR method
- heat to 95 degrees - break H bonds & make DNA single stranded
- cool to 50 degrees - allow primers bind - comp base pairing - dna double stranded - dna polymerase can bind
- heat to 70 degrees - dna polymerase adds comp nucleotides- forms phosphodiester bonds
- repeat - each cycle doubles DNA
1-2-4-8-16
what is a vector
something thats used to move DNA from one place to another
what are virus
bacteriophage
what are plasmids
double stranded loop of DNA that transfers genes between bacteria
how do you insert a gene into a vector
- use same restriction enzyme to cut plasmid
- sticky ends will be comp
- dna ligase reforms phosphodiester bond
- forms recombinant dna (dna from more than one source)
how do you insert vector into bacteria
- ice cold CaCl2
- heat shock
INC PERM OF BACTERIAL CELL WALL
what is a transgenic organism
contains rDNA
what is electrophoresis used for
using electricity to separate DNA fragments by length
electrophoresis
- DNA negatively charged
- attach fluorescent label or stain DNA frag
- put DNA frag into well at negative end
- turn on current
- dna frag attracted to positive electrode
- small frag faster (further)
in electrophoresis how do you calibrate the scale
wiring pieces of dna of known length
3 fragments =
2 cuts
what are marker genes
genes paired with target genes to check if vector inserted properly
give types of marker gene
- UV fluorescence
- antibiotic resistance
what do transformed bacteria contain
rDNA (target gene and marker gene)
only bacteria that have accepted the vector will..
- will fluoresce under UV
- will be able to survive in culture with antibiotic
what do you do once transformed bacteria have been identified?
select and culture transformed bacteria
what is a dna probe
a short sequence of dna with label attached that is complementary to a specific allele/mutation/gene
what are dna probes used for
used to test if sample of dna contains specific sequence
how dna probes work
- attach label to dna probe
- if sequence (disease causing allele) present
- dna probe will hybridise and stick to dna sample with comp base
- rinse to remove unhybridised dna probes
- view labels (uv fluorescene, radioactive) - dna invisible
what must dna be when using dna probes
single stranded
dna microarray
- label attached to human dna (not probe)
- test multiple alleles/mutations at once
- many dna probes attached to tile in grid (know positions)
- attach patients dna with label
- dna will hybridise to comp dna
- rinse —> view labels
uses in agriculture (soya beans, golden rice)
- express protein from bacteria
- protein is toxic to insects
- fewer insects eat soya plant
- gene from corn —> rice
- express vitamin A
advantages of transformed organisms in agriculture
- use less chemical pesticide
- more efficient food chain
- prevents blindness caused by vitamin A deficiency
disadvantages of using transformed organisms in agriculture
- monoculture of crop has low genetic diversity
- susceptible to enviro factors e.g. disease
- have to buy seeds every year
- dec in biodiversity
uses of transformed organisms in industry and research
- making enzymes
- e.g. rennin, lipase
- transformed pathogens to TREAT disease
- attack other pathogens but dont infect humans
advantages of using transformed organisms in industry and research
- reduce energy and cost
- fast and cheap production
- treat disease
- pathogens wont develop resistance
- reduce suffering
disadvantages of using transformed organisms in industry and research
- could mutate and infect humans
- could be used in war
uses of transformed organsism in medicines (insulin)
- transform bacteria to express proteins
- mammals can also be transformed to produce useful prod in their milk
advantages of using transformed organisms in medicine
- makes human proteins
- cheaper and easier than making proteins synthetically
disadvantages of using transformed organisms in medicine
- possible unexpected problems e.g. cancer in animals
- using animals as commodities
what is genetic counselling used for
- identify carriers of genetic disease
- specific allele
- most effective treatment
genetic counselling
(USES OF DNA PROBES)
- healthcare professionals advise people/parents about risks
- passing on inheritable diseases
- developing diseases later in life
- e.g. angelina jolie
- BRCA1- 87% chance of developing breast cancer
- double mastectomy
- make informed decisions about treatments and preventions
genetic screening
(USES OF DNA PROBES)
- parents can see if they are carriers of recessive alleles (IVF)
- can diagnose and treat before symptoms show
- e.g. babies screened for CF
personalised medicine
(USES OF DNA PROBES)
- if doctor knows patients genotype - give best drugs for them
- mutation X - drug X
- mutation Z - drug Z
- change drugs if know patient has allele that causes side effect with normal drug
what is genetic fingerprinting
identifying individuals by comparing difference in Variable Number Tandem Repeats (VNTRs)
what does non-coding dna contain lots of
VNTRs
- dont code for proteins
- dont affect phenotype - vary more than coding dna
- number of repeats of each VNTR varies
relatedness
USES OF GFP
- more closely related two individuals - high percentage match of VNTRs
- conservation of endangered species
- avoid inbreeding by matching individuals who = least related
- maintains genetic diversity
forensic use
USES OF GFP
- samples of dna from crime scene and suspects
- amplify using PCR
- run gel electrophoresis
- chance of 2 individuals having same number of VNTRs very low
medical diagnosis
USES OF GFP
- test for specific combinations of alleles
- used to diagnose disorders
e. g. embryo screening for IVF
