Mouse Design

Question 1

genetic homology between humans and mice

Answer 1

99%

Question 2

physiological similarities between humans and mice

Answer 2

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

Question 3

what vectors are used to transfer genes in transgenic modification?

Answer 3

bacterial plasmid

Question 4

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

Answer 4

male pronuclei of fertilised egg

Question 5

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

Answer 5

pseudo-pregnant females

Question 6

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

Answer 6

DNA nserted at different integration sites

Question 7

components of transgene:

Answer 7

gene of interest, reporter construct, control element

Question 8

how are transgenic lines produced?

Answer 8

founder offspring backcrossed to WT

Question 9

how are offspring screened for transgene?

Answer 9

PCR, southern blot and probe

Question 10

random integration refers to:

Answer 10

each egg possessing different copies at different locations

Question 11

transgene

Answer 11

cDNA or cloned genomic DNA

Question 12

what determines where and when the transgene is expressed?

Answer 12

promoter

Question 13

CMV

Answer 13

strong promoter leading to constitutive expression in mammals

Question 14

difference between CMV and cell-specific promoter

Answer 14

cell-specific promoter allows for regulated gene expression

Question 15

cell-specific promoter

Answer 15

GFAP

Question 16

why is DNA microinjected into the pronuclei of fertilised eggs?

Answer 16

so genes introduced into germ line

Question 17

ALS mouse model

Answer 17

G85R placed into SOD1 and injected into male zygote pronuclei

Question 18

ALS phenotype (4 signs)

Answer 18

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

Question 19

increased endogenous SOD1 expression correlated with

Answer 19

younger onset

Question 20

how do interference RNA knockdown genes

Answer 20

microRNA or shRNA disrupt transcription

Question 21

dominant negative construct

Answer 21

mutant protein impairs function of wild-type protein

Question 22

cre recombinanse

Answer 22

recombination at loxp sites

Question 23

negatives of transgenic over-expression

Answer 23

ectopic expression
multiple copies inserted
insertional mutagenesis

Question 24

how can random integration lead to insertional mutagenesis?

Answer 24

if inserted into exon or essential element

Question 25

what kind of insertion does not affect gene expression?

Answer 25

intergenetic

Question 26

How does transgenic insertion into exons or introns cause confounding phenotypes?

Answer 26

insertional mutagenesis

Question 27

What kind of confounding phenotypes does random integration usually cause?

Answer 27

KO

Question 28

Zfp36 KO models:

Answer 28

arthritis, dermatitis

Question 28

Zfp36 KO models:

Answer 28

arthritis, dermatitis

Question 29

ROSA26

Answer 29

produces mRNA which does not code for a protein

Question 30

KO generation

Answer 30

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

Question 31

how are transgenes incorporated into ES?

Answer 31

electroporation

Question 32

neomycin gene

Answer 32

positive selection (will survive in presence of geneticin)

Question 33

what does survival from geneticin not control against?

Answer 33

random integration

Question 34

how does gancyclovir select against random integration?

Answer 34

converts tK into toxic compound
presence of tk means homologous recombination has not occurred as tk is outside of the arms

Question 35

why do chimeric mice have fur of both colours?

Answer 35

developed from injected and native ES stem cells

Question 36

chimeric mice bred with WT to produce

Answer 36

F2 KO

Question 37

constitutive KO + KI disadvantages

Answer 37

may cause embryonic/perinatal lethality/compensatory upreguation of related genes, Neo gene can alter expression levels (in KI)

Question 38

solution of embryonic/perinatal lethality

Answer 38

cell-type specific expression and inducible knockout

Question 39

solution to compensatory changes

Answer 39

inducible knockout in adulthood

Question 40

how to prevent neo-induced changes to gene expression?

Answer 40

neo removal