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ANBI 470 > M3 L2 > Flashcards

M3 L2 Flashcards

1
Q

embryonic stem cell mediated gene transfer

A

1) grow embryonic stem cells in culture
2) put transgene in
3) allow cells to be transfected via chemicals
4) put into surrogate

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2
Q

what stage are embryonic stem cells found in?

A

blastocyst stage

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3
Q

what stage forms inner cell mass

A

blastocyst

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4
Q

totipotent

A

potential to become any tissue

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5
Q

T or F: blastocysts are totipotent

A

T

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6
Q

transgenic founder

A

offspring that has the transgene in its gametes & can pass them to their offspring

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7
Q

embryonic stem cell mediated gene transfer offspring are ( ), but only 1 will be the ( )

A

chimera, founder

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8
Q

mosaic vs chimera

A

m-different cells that arise from 1 origin
c-different cells derived from more than 1 origin

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8
Q

advantages & disadvantages of embryonic stem cell insertion

A

A- easy, can differentiate into any cell, can study all pathways

D- chimeras result, harder to find embryonic stem cell lines for livestock

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9
Q

epigenetics

A

describes changes in a phenotype that are not coded in the DNA
- effect of the environment

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10
Q

T or F: epigenetics will modify the phenotype & effect how DNA is regulated

A

T

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11
Q

3 factors that epigenetics can change

A

1) how tightly DNA is coiled around histones
2) methylation of DNA
3) imprinting

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12
Q

more methylated DNA will do what in relation to transcription?

A

prevents it

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13
Q

imprinting

A

parent of origin of each chromosome is known via methylation & can be transcribed different if it comes from father vs mother

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14
Q

T or F: epigenetic tags are not heritable because they are not encoded in the DNA

A

F- it is on the DNA, therefore can be heritable

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15
Q

monozygotic twins

A

identical DNA but may differ in phenotype based off environmental factors
- ex. twins separated at birth

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16
Q

what stage has lots of epigenetic factors? Why?

A

fetus stage, b/c cells have differentiated so can have factors here

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17
Q

somatic cells

A

adult cells

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18
Q

somatic cell nucleus cloning

A

1) take egg cells from 1 breed & remove nucleus
2) take somatic cells & grow in plate
3) fuse somatic cells to enucleated egg via electrical pulse
4) implant into surrogate

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19
Q

disadvantages for cloning

A

1) success rate is low
2) very expensive
3) uses lots of eggs
4) not healthy animals overall
5) ethics

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20
Q

finding of cloning

A

differentiated cells could make an entirely new animal

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21
Q

T or F: programming & epigenetics are taken off cells during cloning

A

T

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22
Q

2 reasons for cloning & example

A

possibility for cells from adult individual to repair a disease
- huntingtons

make vulnerable species more prevelant

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23
Q

at what stage of cloning did scientists discover that a disease could possibly be repaired?

A

cell division

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24
Q

pro-nuclear transfer

A

put whole fertilized egg into recipient

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25
Q

nuclear transfer

A

use somatic cells & put into enucleated oocyte

26
Q

3 methods to get a transgenic founder

A

1) pro-nuclear injection
2) embryonic stem cells
3) nuclear transfer

27
Q

2 ways to interfere with a gene’s function & put in a gene

A

1) knock downs
2) knock outs

28
Q

knock down

A

downregulate product
- prevent gene from being transcribed (can be translated)

29
Q

knockout

A

remove at DNA level
if not there = cannot be transcribed OR translated

30
Q

2 types of knockout methods

A

1) homologous recombination
2) non-homologous end-joining

31
Q

what are antisense genes made of

A

single stranded RNA complementary to mRNA

32
Q

antisense gene function

A

1) target gene we want to knockdown using complementary RNA
2) gene sequence is then inverted
3) RNA cannot be made into a protein bc the antisense gene is blocking it
4) chews up the double stranded mRNA

33
Q

2 things antisense genes must do:

A

1) be active in same tissues
2) produce large amounts of RNA

34
Q

challenge of antisense genes

A

hard to use RNA bc it degrades very easily

35
Q

1st discovery of antisense genes was in

A

peutinas

36
Q

flavr savr tomato

A

1st genetically engineered crop approved by FDA
- made antisense RNA for PG enzyme

37
Q

3 uses of antisense RNA in plants

A

1) alteration of flower colour
2) fruit ripening
3) virus resistance

38
Q

example of GMO using an antisense gene

A

flavr savr tomato

39
Q

gene shears

A

targets a specific sequence that you want cut
- ribosome cuts mRNA that it is complementary to = no protein made

40
Q

what are gene shears made of

A

ribosomes (RNA)

41
Q

T or F: gene shears are normally present

A

T, b/c they are ribosomes

42
Q

how are gene shears delivered

A

viral vectors in mammalian cell cultures

43
Q

RNA interference (RNAi)

A

uses microRNA that gets cut into small pieces call small interfering RNAs & take up by protein complexes = binds to target area & cuts = no protein made

44
Q

why are double stranded RNA used for RNA interference?

A

bc single stranded will degrade easily

45
Q

what enzyme cleaves the target mRNA in RNA interference?

A

mRNA

46
Q

morpholino knockdown

A

synthesized oligionucletides that are complementary to a specific target, therefore blocking translation/splice sites

47
Q

2 advantages of morpholinos

A

1) not degraded by nucleases b/c they are synthesized, not natural oligionucleotides
2) do not trigger immune response

48
Q

T or F: morpholinos are incorporated into the genome

A

F

49
Q

how does morpholino knockdown work?

A

1) insert morpholino into target gene using microneedle
2) antisense to targeted DNA
3) blocks translation = no protein produced

50
Q

example of morpholino knockdown

A

zebrafish ocular phenotypes-> were different

51
Q

non-homologous end-joining

A

repair breakage of DNA through error prone pathway that results in a deletion or insertion in target gene, does not require a template

52
Q

2 ways double stranded breaks in the genome can be repaired

A

1) homologous recombination
2) non-homologous end-joining

53
Q

homologous recombination

A

nucleotide sequences are exchanged between 2 similar/identical molecules of DNA during recombination
- insertion of gene/correction
- requires a template

54
Q

2 functions of neomycin

A

1) disrupts gene
2) use as indicator of effectiveness b/c if there is resistance = know transgene is inside cells

55
Q

gene knock out using homologous recombination

A

create a transgene very close to gene we want knocked out & add in neomycin cassette inside transgene

56
Q

neomycin

A

antibiotic resistance gene

57
Q

HSV TK function

A

serves as indicator that transgene is fully incorporated into the genome

58
Q

how many markers are present with 1 homologous recombination event vs 2?

A

1- both markers are still present
2- NEO present, TK lost

59
Q

insert targeting vector into embryonic stem cells

A

put targeting vector into nucleus of cell then rely on homologous recombination so it will flip inside to the genome = cross over piece put into normal genome & gene is released

60
Q

crossover must occur ( ) to ensure gene is successfully knocked out

A

2 crossover events

61
Q

knockout animal example

A

zenotransplantation

62
Q

how does xenotransplantation work

A

pig Gal sugar is not present in humans -> rejection of pig organs due to antibody response
- Gal knockout was developed

ANBI 470 (23 decks)

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