Developmental Exam I

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How can you birthdate a neuron?

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1

How can you birthdate a neuron?

add a molecule that gets added during replication or use antibodies to tag PCNA

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2

tagging PCNA vs molecule replication

PCNA is only good short term and antibodies are species specific and molecule replication used to be radioactive

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3

Lineage Analysis vs Birthdating

birthdating tags new neurons while lineage analysis provides a family tree

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4

How does lineage analysis with a retrovirus work?

a cell is infected with a modified retrovirus which replenishes with division tagging new cells

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5

Pros and Cons of retrovirus

a pro is that it contains a reporter gene but it cannot replicate

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6

Retrovirus Examples

Horseradish peroxidase and B-Galactosidase

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7

How does lineage analysis with Cre-LoxP work?

cre is an enzyme that causes recombination at a floxed DNA site

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8

what are major challenges to studying gene expression?

  • multiple time points to be studied

  • Neuroplasticity

  • diverse cell populations intermingle

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9

what methods can find DNA anatomically?

Immunohistochemistry

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10

What can you learn using immunohistochemistry?

where protein is expressed using primary and secondary antibodies

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11

Pros and cons of immunohistochemistry

it provides spatial information but antibodies have to be made and are species specific

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12

what methods can find DNA molecularly?

Western blot and Elisa

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13

what can you learn using western blot?

the quantity of protein expressed

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14

Pros and Cons of western blot

It comes in a large quantity but gives no spatial info and needs antibodies

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15

How is Elisa used?

there is a 96 well plate lined with antibodies where proteins are read

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16

What method can find mRNA anatomically?

In Situ Hybridization

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17

what does ISH tell you?

where mRNA is expressed

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18

Pros and Cons of In Situ Hybridization

It provides spatial info and has a wide range of probes but the probes are specific

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19

What methods can find mRNA molecularly?

Northern Blot and qRT-PCR

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20

What is northern blot used for?

to figure out if mRNA is present or not

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21

Pros and Cons of Northern Blot

the cost is low and its fast, but there is no spatial info

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22

what does qRT-PCR tell you

the specific quantity of mRNA

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23

Pros and cons of qRT-PCR

it is fast and can measure a lot but there is no spatial info and it is hard to be specific

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24

what is a primary antibody?

It finds a proteins

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25

what is a secondary antibody?

it finds the primary and reveals it

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26

Carnegie Stage 1

fertilization of the egg

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27

Carnegie Stage 2

first cleavage through morula

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28

Carnegie Stage 3

unimplanted blastocyte

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29

what does the inner cell mass become?

the embryo

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30

what does the outer cell mass become?

the placenta

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31

when do the inner and outer cell mass form?

Carnegie Stage 3

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32

Carnegie Stage 4

shedding of zona pellucida

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33

Carnegie Stage 5

implantation is complete

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34

Carnegie Stage 6

Gastrulation begins and the primitive steak appears

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35

When do the 3 germ layers form?

Carnegie Stage 6

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36

What are the 3 germ layers?

endoderm, mesoderm, ectoderm

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37

Carnegie Stage 7

cell migration defining gastrulation

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38

what does the endoderm turn into?

insides and organs

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39

what does the mesoderm turn into?

muscles

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40

what does the ectoderm turn into?

skin and nervous system

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41

Carnegie Stage 8

appearance of neural plate and neural groove

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42

Carnegie Stage 9

somites appear on either side of the neural groove

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what are somites?

precursors of dermal tissue, muscles, vertebrae

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Carnegie Stage 10

formation of neural tube

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45

Carnegie Stage 11

more somites grow

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46

Carnegie Stage 12

neural tube closes

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47

Carnegie Stage 13

30 pair of somites

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48

Carnegie Stage 14

rapid brain growth with definition of forebrain, hindbrain, and midbrain

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49

Carnegie Stage 15

cerebral hemispheres can be discerned

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50

what are the three primary brain vesicles?

prosencephalon, mesencephalon, and rhombencephalon

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51

which of the primary brain vesicles subdivide further?

prosencephalon and rhombencephalon

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52

what does the prosencephalon divide into?

the telencephalon and diencephalon

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53

what does the rhombencephalon divide into?

metencephalon and myelencephalon

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54

What does the telencephalon form?

cerebrum

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55

what does the diencephalon form?

eye cups and thalamus

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56

what does the metencephalon form?

pons and cerebrum

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57

what does the mesencephalon form?

midbrain

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58

what does the myelencephalon form?

medulla oblongata

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59

What defects are caused by the failure to close the anterior/cranial neuropore?

iniencephaly, encephalocele, and anencephaly

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60

characteristics of iiencephaly?

“stargazing” posture with a stillbirth or premature death

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Characteristics of encephalocele?

herniated meninges and brain tissue

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62

Characteristics of anencephaly

total or partial absence of brain

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63

What failed to close in craniorachischisis?

middle portion

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Characteristics of craniorachischisis?

anencephaly along with lesion on spinal cord with “stargazing” look

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what failed to close in spina bifida?

posterior/caudal neuropore

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Characteristics of spina bifida

herniation of meninges and spinal cord

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mutation

change in DNA sequence

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68

null mutation

complete failure to express protein coded by mutated gene

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69

hypomorph

wild type gene expressed at a low level

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70

wild type

standard, unmutated gene

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strain

a reproductively isolated group of animals

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72

inbred strain

over 20 consecutive generations of sibling mating to produce homozygosity

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73

isogenic

strain where every individual is genetically identical

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74

What is forward genetics?

identifying a gene then finding the phenotype

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75

What is reverse genetics?

selecting a gene to then find a phenotype

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76

Forward vs Reverse genetics

forward genetics has no bias of previous knowledge and can exploit spontaneous mutations while reverse genetics must inactivate a known gene and the mutation may not be visible

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77

What committee oversees institutional use of animals

Institutional Animal Care and Use Committees (IACUC

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78

Theiler Stage 1

fertilization

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Theiler Stage 7

blastocyst sheds zona pellucida and implants

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Theiler Stage 11

appearance of neural plate and neural groove

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Theiler Stage 12

portion of neural tube closes

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Theiler Stage 14

anterior portion of tube closes

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83

Theiler Stage 15

posterior neuropore forms

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Theiler Stage 16

posterior neuropore closes

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Theiler Stage 18

rapid brain growth

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86

Danio rerio stage 1

early blastula

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87

Danio rerio stage 2

midblastula

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Danio rerio stage 3

early gastrula

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Danio rerio stage 4

midgastrula

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Danio rerio stage 5

early segmentation stage

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91

Danio rerio stage 6

mid-segmentation stage

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92

Danio rerio stage 7

pharyngula stage

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93

Drosophila melanogaster formation

embryo → 1st instar → 2nd instar → 3rd instar → prepupa → pupa

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94

Bownes Stage 1-3

unusual pattern of cleavage

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95

Bownes Stage 4

syncytial blastoderm

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96

Bownes Stage 5

cellular blastoderm

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Bownes Stage 6

gastrulation

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98

Bownes Stage 7

germ band elongates

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99

Bownes Stage 9

neuroblasts form

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100

Bownes Stage 11

delamindation of neuroblasts

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