Mouse Design Flashcards

1
Q

genetic homology between humans and mice

A

99%

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2
Q

physiological similarities between humans and mice

A

cerebellum, cortex, thalamus, medulla, pons and olfactory bulb

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3
Q

what vectors are used to transfer genes in transgenic modification?

A

bacterial plasmid

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4
Q

where is the gene of the interest transferred during random integration?

A

male pronuclei of fertilised egg

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5
Q

once the DNA is randomly inserted into genome of pronuclei, where are the eggs implanted?

A

pseudo-pregnant females

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6
Q

why do founder lines differ despite being infected with same DNA?

A

DNA nserted at different integration sites

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7
Q

components of transgene:

A

gene of interest, reporter construct, control element

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8
Q

how are transgenic lines produced?

A

founder offspring backcrossed to WT

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9
Q

how are offspring screened for transgene?

A

PCR, southern blot and probe

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10
Q

random integration refers to:

A

each egg possessing different copies at different locations

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11
Q

transgene

A

cDNA or cloned genomic DNA

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12
Q

what determines where and when the transgene is expressed?

A

promoter

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13
Q

CMV

A

strong promoter leading to constitutive expression in mammals

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14
Q

difference between CMV and cell-specific promoter

A

cell-specific promoter allows for regulated gene expression

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15
Q

cell-specific promoter

A

GFAP

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16
Q

why is DNA microinjected into the pronuclei of fertilised eggs?

A

so genes introduced into germ line

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17
Q

ALS mouse model

A

G85R placed into SOD1 and injected into male zygote pronuclei

18
Q

ALS phenotype (4 signs)

A

poor righting and hindpaw grasping
paralysis 2 weeks after clinical onset
aberrant hindlimb posture within 3 days of clinical onset

19
Q

increased endogenous SOD1 expression correlated with

A

younger onset

20
Q

how do interference RNA knockdown genes

A

microRNA or shRNA disrupt transcription

21
Q

dominant negative construct

A

mutant protein impairs function of wild-type protein

22
Q

cre recombinanse

A

recombination at loxp sites

23
Q

negatives of transgenic over-expression

A

ectopic expression
multiple copies inserted
insertional mutagenesis

24
Q

how can random integration lead to insertional mutagenesis?

A

if inserted into exon or essential element

25
what kind of insertion does not affect gene expression?
intergenetic
26
How does transgenic insertion into exons or introns cause confounding phenotypes?
insertional mutagenesis
27
What kind of confounding phenotypes does random integration usually cause?
KO
28
Zfp36 KO models:
arthritis, dermatitis
28
Zfp36 KO models:
arthritis, dermatitis
29
ROSA26
produces mRNA which does not code for a protein
30
KO generation
homologous recombination to produce the target gene in ES pos and negative selection of ES stem cell implantation into blastocyst and injection into foster mother
31
how are transgenes incorporated into ES?
electroporation
32
neomycin gene
positive selection (will survive in presence of geneticin)
33
what does survival from geneticin not control against?
random integration
34
how does gancyclovir select against random integration?
converts tK into toxic compound presence of tk means homologous recombination has not occurred as tk is outside of the arms
35
why do chimeric mice have fur of both colours?
developed from injected and native ES stem cells
36
chimeric mice bred with WT to produce
F2 KO
37
constitutive KO + KI disadvantages
may cause embryonic/perinatal lethality/compensatory upreguation of related genes, Neo gene can alter expression levels (in KI)
38
solution of embryonic/perinatal lethality
cell-type specific expression and inducible knockout
39
solution to compensatory changes
inducible knockout in adulthood
40
how to prevent neo-induced changes to gene expression?
neo removal