Notes 3 and 4

Question 1

Southern blotting steps

Answer 1

1. DNA run onto gel: looks like smear
2. Filter paper holds DNA
3. Use probe thats highly specific to your gene of interest
4. Wash away unbound probe
5. Develop autoradiogram and expose to xray film
6. Capture image on film

Question 2

Type of filter paper used in Southern bloting

Answer 2

nitrocellulose

Question 3

In Situ Hybridization (4 steps)

Answer 3

1. Gently denature chromosomes that are in metaphase
2. Use probe labeled with fluorescent dye
3. Denature and hybridize
4. Locate where it is on chromosome “the site”

Question 4

Northern blotting

Answer 4

Transfer of RNA from a gel to a membrane

Question 5

Western blotting steps

Answer 5

1. Separate proteins on SDS gel
2. Transfer to nitrocellulose membrane using electrotransfer
3. This is called: Polyacrylamide gel electrophoresis
4. Use antibodies to give specificity to proteins

Question 6

What does SDS do?

Answer 6

coats proteins with a negative charge

Question 7

How are proteins separated by polyacrylamide gel electrophoresis

Answer 7

Separated on size

No shape or charge component

Question 8

Yeast model organisms

Answer 8

Saccharomyces cerevisiae

Schizosaccharomyces pombe

Question 9

Budding yeast

Answer 9

Saccharomyces cerevisiae

Question 10

Fission yeast

Answer 10

Schizosaccharomyces pombe

Question 11

Slime Mold

Answer 11

Dictyostelium discoideum

Question 12

Nematodes

Answer 12

Caenorhabditis elegans

Question 13

Fruit fly

Answer 13

Drosophila melanogaster

Question 14

Silk worm

Answer 14

Bombyx morii

Question 15

Frogs

Answer 15

Rana pipens

Xenopus laevis

Question 16

Typical dissection frog

Answer 16

Rana Pipens

Question 17

African clawed frog

Answer 17

Xenopus laevis

Question 18

Zebrafish

Answer 18

Danio rerio

Question 19

Mouse

Answer 19

Mus musculus

Question 20

Vascular plants

Answer 20

Arabidopsis thaliana

Question 21

What’s special about wood frogs?

Answer 21

They’re very freeze tolerant and will freeze solid in winter

Question 22

How do they prevent ice from damaging them?

Answer 22

Have polysaccharides to make sure ice doesn’t damage them

Question 23

when the sperm and egg fuse its called

Answer 23

syngamy

Question 24

When does the first cleavage occur?

Answer 24

90 minutes

Question 25

When does the gastrula occur?

Answer 25

10 hours | 20,000 cells

Question 26

When does division slow (cell number)

Answer 26

4,096 cells

Question 27

Upper side of the egg is called

Answer 27

Animal hemisphere

Question 28

Lower side of the egg is called

Answer 28

Vegetal hemisphere

Question 29

When it divides to four cells, those cells are called...

Answer 29

blastomeres

Question 30

At 4,000 cells there's the ______ stage

Answer 30

Blastula

Question 31

What forms inside the blastula

Answer 31

blastocoel

Question 32

What forms after the blastula?

Answer 32

Gastrula

Question 33

When does the notochord and neurula form?

Answer 33

about 13 hours

Question 34

When does a tail bud form?

Answer 34

1 day

Question 35

When does a tadpole form

Answer 35

3 days

Question 36

What is the mid blastula transition?

Answer 36

Where RNA synthesis occus and germ layers can develop

Question 37

Mesoderm

Answer 37

muscle, bone

Question 38

Ectoderm

Answer 38

skin, nervous system

Question 39

Endoderm

Answer 39

gastro lining

Question 40

How is the frog cell cycle abbreviated?

Answer 40

only has S & M stage | Replicate, divide

Question 41

Rapidly proliferating cells

Answer 41

Blastomeres

Question 42

5 questions of inquiry

Answer 42

1. Observation and questions 2. hypothesis 3. testing the hypothesis 4. Results 5. Conclussion

Question 43

Steps of flow cytometry

Answer 43

1. Prepare embryonic nuclei from blastula 2. Stain with propidium iodide 3. excite with UV 4. Quantify fluorescence by flow cytometry to determine DNA content

Question 44

How is the mesoderm formed?

Answer 44

Contacting animal and vegetal layers

Question 45

How do we break down cell layer?

Answer 45

CMFM (calcium magnesium free medium)

Question 46

What are dispersed cells?

Answer 46

They aren't touching

Question 47

What are dissociated cells?

Answer 47

They're touching, in a pile

Question 48

What happened with dispersed cells vs. dissociated?

Answer 48

Dispersed cells lost their way and only stayed in S phase. Dissociated were similar to an intact embryo and still developed.