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BB10006 Cell and Molecular Biology > Animal Development (excluding development of Xenopus, Drosophila and Mouse) > Flashcards

Animal Development (excluding development of Xenopus, Drosophila and Mouse) Flashcards

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1
Q

What makes a good animal model?

A

any species that’s been widely studied, easy to maintain, easy to breed, low cost, known genome sequence, experimental advantages.

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2
Q

What would make an animal model good for genetics studies?

A
  • large array of mutants
  • large no. of offspring
  • short generation time
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3
Q

What would make an animal good for embryology?

A
  • robust embryos
  • easily manipulated
  • large no. of embryos
  • external development
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4
Q

What would make an animal good for genomics?

A
  • relevance to human genome
  • disease model
  • drug testing
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5
Q

Examples of developmental models (invertebrates)

A
  • CAENORHABDITIS ELEGANS - worm

- DROSOPHILIA MELANOGASTER - fly

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6
Q

Examples of Developmental models (vertebrates)

A
  • DANIO RERIO - zebrafish
  • XENOPUS LAEVIS - African clawed frog
  • GALLUS GALLUS DOMESTICUS - chicken
  • MUS MUSCULUS - mouse
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7
Q

Definition of Blastula stage

A

ball of cells

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8
Q

Definition of Gastrulation

A

cell movement producing germ layers

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9
Q

Definition of germ layers

A

ectoderm, mesoderm, endoderm

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10
Q

Definition of neural plate stage

A

embryo after gastrulation

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11
Q

Definition of neurulation

A

cells move to roll up neural tube

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12
Q

Definition of dorsal and ventral

A

back and belly

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13
Q

Steps of animal cloning (somatic cell nuclear transfer)

A

1) UV radiation of unfertilised egg
2) nucleus of cell of blastula taken and injected into enucleated egg
3) albino frog

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14
Q

Steps of cloning with differentiated adult cells

A

1) Cultured adult skin cells
2) Uv radiation destroys DNA in nucleus of egg
3) removal of nucleus from cultured cells
4) transfer of nucleus to egg
5) tadpole develops

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15
Q

What are transcription factor codes in development?

A
  • bind to DNA to cause transcription of genes, lots can be active at same time
  • If diff transcription factors activated later on in development, it may differentiate differently and differ from wild type.
    Wild type of is blank / TF1 / TF1 + 2
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16
Q

What are inductive signals and what is their role in development?

A
  • signal transduction carries signal from membrane to nucleus
  • cause activation of diff TF
  • signalling cell manufactures inductive signals
  • further away you get lower conc of IS
  • receptors span plasma membrane, signal binds, protein enters nucleus and activates trans factor
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17
Q

What dose of Inductive signal does TF1 and TF2 need?

A
  • TF1 is medium
  • TF2 is high
  • low dose = no TF switched on
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18
Q

What is experimental embryology (using signal cells)

A
  • taking and moving signal cells to other end of cells to see what would happen , high dose at each end
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19
Q

Why are amphibians well suited to experimental embryology? (stage 1 of understanding inductive signals)

A
  • mother produces lots of eggs
  • embryos develop externally
  • large embryos
  • tissue heals well
  • embryos wont die if taken elsewhere
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20
Q

What did Hans Spearmann do in his experiment and what were the aims?

A
  • carried out experiment in Newts in 1900’s
  • took fertilised eggs, wanted to split in half but not separate them
  • wrapped hair around an egg to keep cytoplasm on one side and nucleus on other
  • one cell would go to the other side so he got two early embryos occurring
  • only one side had grey crescent, other side develops oddly
21
Q

What occurs in a Spearmann Organiser Graft?

A
  • Gastrala stage embryo –> cut out a piece of tissue at dorsal region where dorsal lip would occur
  • transferred to ventral half of gastrin stage embryo
  • allowed embryos to develop
  • got two headed twins
  • THEREFORE Spearmann Organiser Graft produces all signals you need to induce second axis
22
Q

What is an inductive signal?

A

Extracellular signal which causes cells to follow different fates.

23
Q

What is cell fate?

A
  • what cells will end up being due to differentation
24
Q

What changes in fate does a Spearmann Organiser Graft make?

A
  • induces dorsal fate in cells which would otherwise follow ventral
  • cells in developing embryo that induces development or central nervous system
25
Q

What is the role of a signal transduction?

A
  • Relays the message from the extracellular signal from the membrane to the nucleus where it activate gene expression e.g Transcription Factor
26
Q

What occurs in stage 2 (Generating Mutants) of understanding inductive signals?

A
  • make plasmid library containing 1000’s of random genes
  • use a needle to inject mRNA into embryo
  • lots of protein translated by embryo
  • abnormal embryo / tadpole due to overexpression
27
Q

What happens if fertilised egg treated with UV radiation?

A
  • forms into belly piece embryos

- so lots of diff mRNA’s injected to find one with resolves the head development

28
Q

What does noggin do?

A
  • promote dorsal structures
  • it’s a secreted signalling molecule
  • it’s expressed in the Spearmann Organiser - cells far away follow ventral fate, cells close follow dorsal fate
  • doesn’t turn on the HOX gene
29
Q

What occurs in stage 3 (checking whether gene is expressed) In-situ hybridisation of understanding inductive signals?

A

1) Add RNA probe attached to tag
2) Add enzyme which recognises tag, enzymes are attached to antibodies
3) Add substrate to solution to form a coloured precipitate

30
Q

What is superficial cleavage in Drosophila?

A
  • divides as nuclei not cells, goes from one nuclei to 2 to 4 etc.
  • nuclei increase
  • after 9th division nuclei moves to outside edges of zygote, called syncytial blastoderm
  • pole cells move to end of zygote
31
Q

What is a genetic screen and what are the stages to it?

A
  • technique used to identify and study an interesting phenotype within a mutated population.
    1) Generate the mutants
    2) Identify gene that has mutated
    3) Find where genes expressed using forward genetics
32
Q

What occurs in stage one of a genetic screen (generating the mutants)?

A
  • F1 generation leads to recessive phenotype, not clear which individuals have mutated gene though
  • so gene crossed with wild type to make F2 generation
  • F2 then inbred and from this you can see which families contain the mutated gene (they will have inherited 2 copies)
33
Q

What are homeotic mutants?

A
  • expressed in diff anterior/posterior positions
34
Q

What are homeotic genes?

A
  • Homeobox transcription factors
  • activate/repress genes required to specify organs/cell types
  • control homeobox domain
  • control pattern of body formation
35
Q

What is collinearity?

A
  • order of expression in embryo matches order of genes
36
Q

Drosophila MOST OF HEAD (position in body related to homeotic gene wildtype)

A

lab and Dfd give antennae

37
Q

Drosophila Thoracic seg 1 (position in body related to homeotic gene wildtype)

A

scr gives pair of legs

38
Q

Drosophila Thoracic seg 2 (position in body related to homeotic gene wildtype)

A

Antp gives pair of legs and wings

39
Q

Drosophila Thoracic seg 3 (position in body related to homeotic gene wildtype)

A

Ubx and Antp give pair of legs and halteres

40
Q

Drosophila Abdominal seg 1-4 (position in body related to homeotic gene wildtype)

A

abdA and Ubx and Antp give no outgrowths

41
Q

What are the homeobox genes like found in mammals compared to Drosophila?

A
  • 38 hox genes, similar
42
Q

What does lineage restriction mean?

A

cell will only give rise to certain cell types

43
Q

Issues with doing a genetic screen on a mouse

A

need something cheaper smaller and easier to breed

44
Q

What is reverse genetics?

A

add/manipulate gene then look at phenotype

- permanent change

45
Q

Transgenics in a mouse (overexpressing gene product)

A
  • male pronuclei larger
  • use needle to inject DNA into male pronuclei
  • use fungi plasmifs in bacteria (replicate), use promotors in a test tube and DNA polymerase to drive transcription –> lots of copies of gene
  • DNA will intergrate into chromosomes
  • done in dish, inject eggs into mother
46
Q

What is a targeting construct?

A
  • knock out genes
  • made up of DNA stitched together, sequences contained in plasmids
  • purified from bacterial plasmids, linearized and used within ES to cause genetic deletion
  • DNA construct moves into ES nucleus, recombination
  • gene of interest removed and replaced with a marker and reporter
  • ES cells reseeded into culture dish- grow
47
Q

What is a chimeric mouse?

A
  • combo of normal and targeted cells
48
Q

Microinjection of ES cells into host blastocyst

A
  • take blastocyst embryos, inject into cavity of blastocyst and expel out the ES cells
  • get recombinant blastocysts
  • transplant into uterine horn not oviduct of surrogate mother

BB10006 Cell and Molecular Biology (5 decks)

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