CMMB 403 final exam actual questions and answers 2024-2025 with complete solution
what does CRISPR use in order to generate changes in genome? is CRISPR forward or
reverse genetics? Ans- - nuclease
- 20 nucleotide RNA sequence to target specific sites in the genome
- reverse genetics
what is the role of Cas9 in CRISPR Ans- endonuclease that catalyzes double stranded
breaks
what are the two repair paths cells can use to repair double stranded breaks from CRISPR-Cas9 Ans- 1. non-homologous end joining (NHEJ)
- naturally try to repair
- imperfect, creates indels causing frameshift mutation
2. directed homology repair
- sequence homology on 5' and 3' ends to encourage homologous recombination flanking the double stranded breaks
- insert known sequences to try and treat genetic diseases
explain the method of conditional gene inactivation Ans- - make targetting construct in lab
- electroporate construct into embryonic stem cells to target endogenous Shox2 locus
- mice generated by blastocyst injection into ESCs carry loxP sites therefore can delete Shox2
2 components for a conditional knockout mouse Ans- 1. transgenic mice express Cre recombinase in the tissue of interest
2. mice with loxP sites on each side of the gene or region to be deleted
How is CRISPR-Cas9 being used in a patient in the USA to treat sickle cell anemia? Ans- by inactivating an enhancer of the BCL11A gene
(BCL11A gene represses fetal hemoglobin)
how does the BCL11A protein affect global gene expression Ans- it represses fetal hemoglobin gene expression
In what kind of animals is CRISPR good? Ans- any animals for which one cell embryos
are accessible - good for mammals, not chickens
what is the result of CRISPR inactivating Rspo2 in half a frog? Ans- Rspo knockout= limbless
side inactivated by CRISPR had no limb
what are transgenic mice good for?
- how are they made?
- is this targeted?
- what are they not useful for? Ans- good for mis-expressing genes, enhancer reporters, etc.
- generated by pronuclear injection; DNA construct is incorporated randomly into the genome
- not useful for LOF because not targeted
how are knockout mice made?
- is it targeted?
- what kind of species is this successful in? Ans- created by genomic targeting in embryonic stem cells
- DNA construct is targeted to specific sites in the genome by homologous recbomination
- mice then made from pluripotent ES cells
- only successful in mice
what is CRISPR/Cas9 good for?
- what kind of species is it successful in? Ans- efficient RNA-directed targeting in single-cell embryos or cell lines from a broad variety of species
- useful for knockout, introducing changes in genome, corrections, etc. - many species
when does organogenesis occur in human development? Ans- 4th-8th week
what day is the neural plate formed? Ans- day 18
what day does the primitive streak disappear? Ans- day 25
what is responsible for neural induction? where are these signals from? Ans- by BMP inhibitors from the pharyngeal endoderm, prechordal plate and notochord mesoderm
on what day does the neural plate consist of columnar cells that are taller than surrounding cells? Ans- day 20-21
during what week does the embryo fold into a 3D structure, most organs start to form, heart starts beating and neural tube forms? Ans- 4th week
- germ disc starts to look more like a body on what day does the neural tube close? Ans- day 28
anencephaly Ans- caused by failure of the anterior neuropore to close
- fatal
craniorachischisis Ans- failure of the entire neural tube to close
- fatal
spina bifida Ans- caused by failure of the posterior neuropore to close
- can be surgically fixed
what are the 3 major divisions of the ectoderm germ layer? Ans- 1. surface ectoderm (epidermis)
2. neural crest 3. neural plate/neural tube
T or F: all 3 major subdivisions of the ectoderm can form neurons Ans- true
what structures does the surface ectoderm form? Ans- - epidermis
- sensory epithelium
- lens/cornea
- etc
what structures does the neural crest form? Ans- - peripheral nervous system (PNS)
- melanocytes
- facial cartilage
- adrenal medulla
- Schwann cells
- glial cells
- sympathetic nervous system
- parasympathetic nervous system
what structures does neural plate/neural tube form? Ans- - brain
- spinal cord
- motor neurons
- retina
which ectoderm subdivisions have high BMPs? low? Ans- surface ectoderm has high BMP levels
neural plate/neural tube has low BMP levels, Sox TFs expressed
define neural plate Ans- portion of the ectoderm which will form the brain and spinal cord
define neurulation
1. primary
what does CRISPR use in order to generate changes in genome? is CRISPR forward or
reverse genetics? Ans- - nuclease
- 20 nucleotide RNA sequence to target specific sites in the genome
- reverse genetics
what is the role of Cas9 in CRISPR Ans- endonuclease that catalyzes double stranded
breaks
what are the two repair paths cells can use to repair double stranded breaks from CRISPR-Cas9 Ans- 1. non-homologous end joining (NHEJ)
- naturally try to repair
- imperfect, creates indels causing frameshift mutation
2. directed homology repair
- sequence homology on 5' and 3' ends to encourage homologous recombination flanking the double stranded breaks
- insert known sequences to try and treat genetic diseases
explain the method of conditional gene inactivation Ans- - make targetting construct in lab
- electroporate construct into embryonic stem cells to target endogenous Shox2 locus
- mice generated by blastocyst injection into ESCs carry loxP sites therefore can delete Shox2
2 components for a conditional knockout mouse Ans- 1. transgenic mice express Cre recombinase in the tissue of interest
2. mice with loxP sites on each side of the gene or region to be deleted
How is CRISPR-Cas9 being used in a patient in the USA to treat sickle cell anemia? Ans- by inactivating an enhancer of the BCL11A gene
(BCL11A gene represses fetal hemoglobin)
how does the BCL11A protein affect global gene expression Ans- it represses fetal hemoglobin gene expression
In what kind of animals is CRISPR good? Ans- any animals for which one cell embryos
are accessible - good for mammals, not chickens
what is the result of CRISPR inactivating Rspo2 in half a frog? Ans- Rspo knockout= limbless
side inactivated by CRISPR had no limb
what are transgenic mice good for?
- how are they made?
- is this targeted?
- what are they not useful for? Ans- good for mis-expressing genes, enhancer reporters, etc.
- generated by pronuclear injection; DNA construct is incorporated randomly into the genome
- not useful for LOF because not targeted
how are knockout mice made?
- is it targeted?
- what kind of species is this successful in? Ans- created by genomic targeting in embryonic stem cells
- DNA construct is targeted to specific sites in the genome by homologous recbomination
- mice then made from pluripotent ES cells
- only successful in mice
what is CRISPR/Cas9 good for?
- what kind of species is it successful in? Ans- efficient RNA-directed targeting in single-cell embryos or cell lines from a broad variety of species
- useful for knockout, introducing changes in genome, corrections, etc. - many species
when does organogenesis occur in human development? Ans- 4th-8th week
what day is the neural plate formed? Ans- day 18
what day does the primitive streak disappear? Ans- day 25
what is responsible for neural induction? where are these signals from? Ans- by BMP inhibitors from the pharyngeal endoderm, prechordal plate and notochord mesoderm
on what day does the neural plate consist of columnar cells that are taller than surrounding cells? Ans- day 20-21
during what week does the embryo fold into a 3D structure, most organs start to form, heart starts beating and neural tube forms? Ans- 4th week
- germ disc starts to look more like a body on what day does the neural tube close? Ans- day 28
anencephaly Ans- caused by failure of the anterior neuropore to close
- fatal
craniorachischisis Ans- failure of the entire neural tube to close
- fatal
spina bifida Ans- caused by failure of the posterior neuropore to close
- can be surgically fixed
what are the 3 major divisions of the ectoderm germ layer? Ans- 1. surface ectoderm (epidermis)
2. neural crest 3. neural plate/neural tube
T or F: all 3 major subdivisions of the ectoderm can form neurons Ans- true
what structures does the surface ectoderm form? Ans- - epidermis
- sensory epithelium
- lens/cornea
- etc
what structures does the neural crest form? Ans- - peripheral nervous system (PNS)
- melanocytes
- facial cartilage
- adrenal medulla
- Schwann cells
- glial cells
- sympathetic nervous system
- parasympathetic nervous system
what structures does neural plate/neural tube form? Ans- - brain
- spinal cord
- motor neurons
- retina
which ectoderm subdivisions have high BMPs? low? Ans- surface ectoderm has high BMP levels
neural plate/neural tube has low BMP levels, Sox TFs expressed
define neural plate Ans- portion of the ectoderm which will form the brain and spinal cord
define neurulation
1. primary
