COCKBURN 1 Flashcards
what is a model organism
a non human species that is used to study a specific phenomenon or disease
why do we use model organisms?
we cannot experiment on humans so we need to learn from other organisms that mimic specific aspects of human biology and are comparatively easy to work with
can be used for forward and reverse genetics
what is forward genetics
there is a phenotype, an observable characteristic that is different, and we figure out which genes regulate those differences
what is reverse genetics
mutate a gene/multiple genes and observe the effect of that on the phenotype
what are the eukaryotic organisms most commonly used?
S.cerevisiae (yeast)
C.elegans
d.melanogaster (fruit fly)
danio rerio (zebrafish)
mus musculus (mouse mouse)
put in order of decreasing ease to grow and increasing similarity to humans
what are characteristics of s.cerevisiae
eukaryotic, unicellular fungus
generation time; 2-3 hours (till next generation is reproductively able)
can exist as haploid or diploid
can reproduce sexually or asexually
can be frozen and revived
what is the life cycle of s.cerevisiae
can be haploid or diploid
haploid yeast can do asexual reproduction by budding
alpha and a are the 2 mating types
can mate to make a diploid cell, which can also do budding
when conditions are hard diploid cell can become haploid by meiosis
what are the advantages of using haploid yeast?
only one copy of each gene, easy to see what happens when you change it
what are characteristics of C.elegans?
invertebrate animal, multicellular
generation time: 3 days, can make 300 progeny
extremely simple, translucent (easy to see inside)
can trance the fate of each cell (1090 total)
two sexes: male and hermaphrodite
can be frozen and revived
how do the two sexes of c.elegans reproduce?
hermaphrodites can self fertilise to make clones of themselves
can also breed with males to make mostly hermaphrodites, but also some males
what is the life cycle of c.elegans
embryo goes through multiple larval stages (L1, L2, L3, L4) till it is a reproductive adult (this all takes about 3 days), and then it ages
an alternative life cycle is if there is crowding starvation or high temperatures at the L1 stage it can go into a dauer state (smaller) and then after several months it can go to L4 and then grow to an adult
what are the characteristics of d.melanogaster
invertebrate animal, multicellular
similar to humans in terms of body organisation, digestive tract, nervous system, excretion, muscles, circulation, skeleton
generation time: 10 days, 100 progeny
share 75% of human disease causing genes
very well studied
what are the characteristics of danio rerio
vertebrate animal, multicellular
generation time: 2-3 months, 200 eggs
optically translucent embryos and larvae (can be seen with light microscope)
relatively simple and inexpensive to maintain, but you need a whole water system
easily treated with small molecules for drug and toxicity screens because it is all in water
what are the characteristics for mus musculus?
vertebrate animal
generation time: 3 months, 2-12 pups
small, easy to house
commonly used to study human biology and perform preclinical testing
not always the perfect models
why do we study unique phenomena
study these unique mechanisms to then maybe apply their unique phenomena to humans
what are some emerging organisms that are being studied for their unique features?
axolotls can regenerate limbs
planaria can regenerate everything, no matter where they are cut
what are the basics of forward genetic screens
perturb lots of genes (randomly or systematically)
look for a specific phenotype
figure out which gene you mutated
how do temperature sensitive mutants work?
permissive temperatures give normal polypeptide folding and the wild type phenotype
restrictive temperatures lead to unfolding/misfolding and proteolysis or aggregation, decreased amount of active protein and yields to temperature sensitive (TS) phenotype
how is replica plating done to isolate TS mutants?
mutate the yeast
put them on a master plate
put it on sterile velveteen
velveteen with imprint of all colonies
make one plate on high temp and one plate on low temp
at low temp all colonies grow
at high temps mutates do not grow, you can go back to the plate and figure out which plates those were
what were the conclusions of mutating cdc1
cdc 1 mutants arrest in G1 phase
cdc1 encodes for a cell wall which is needed to expand
what are the findings of mutating cdc2
arrest in S phase
cdc2 codes for part of DNA polymerase 3
what are the findings of mutating cdc3
stop in mitosis
divide but do not separate, no cytokinesis
cdc3 codes for septin, involved in cytokinesis
what can the fact that yeast can go from haploid to diploid be used for?
can be used to perform complementation
used to test whether two mutants strains have mutations in the same gene
how is a yeast genomic DNA library made?
yeast double stranded DNA is cleaved with restriction nucleases to make thousands of genomic DNA fragments
DNA fragments are inserted into plasmids to make recombinant DNA
introduce into bacteria to make the library
how is the mutant gene identified
introduce different plasmids into yeast and test which plasmids rescue the mutant
sequence the plasmids that rescue to identify the yeast gene responsible for a phenotype
how is the mutant gene identified when investigating human genes
introduce different cDNAs into yeast, test which plasmids rescue the mutant
sequence cDNAs that rescue to identify the human gene responsible for a phenotype
what type of embryogenesis do c.elegans show?
invariant embryogenesis
what does invariant embryogenesis mean for c.elegans?
every embryo develops with the same pattern of divisions and migrations
adult worms have exactly 959 cells generated in the exact same way
very useful when performing genetic screens to understand development
how many cells form during c.elegans embryogenesis and how many die?
1090 cells form
131 die during embryogenesis, leaving 959 cells at adult age
