Lecture 23 Flashcards
Introducing genes to treat diseases
gene therapy
Germ-line gene therapy: genes are introduced into the germ cells to – future generations.
genetically alter
Somatic cell gene therapy: normal genes are added to the – of at least some of the body’s cells to ameliorate the effects of a defective gene
nuclei
Some Goals of Gene Therapy:
Replace – genes with normal genes to cure diseases
defective
Some Goals of Gene Therapy: introduce genes which will enhance –
immune defenses
Some Goals of Gene Therapy: introduce – into cancer cells
suicide genes
Some Goals of Gene Therapy: alter – of genes in the cell
regulation
Some Goals of Gene Therapy: stop the replication of –
viruses
Genes can be introduced to cells in culture (ex vivo) or –
in the patient (in vivo)
+ Adenovirus
Infects many cell types
- Adenovirus
Does not integrate into host genome and can be lost.
+ Adeno-associated virus (AAV)
Integrates into host genome at specific site and cannot be lost
- Adeno-associated virus (AAV)
difficult to work with
- Simple retrovirus
integrate near genes
+ complex retrovirus (lentivirus)
Integrates into host genome and cannot be lost
Integration is not random – occurs away from genes
+ Herpes Simplex Virus (HSV)
DNA stays in nucleus without integrating into genome
- Herpes Simplex Virus (HSV)
only infects nervous system cells
Non-viral methods of gene transfer
liposome/lipolpex or naked DNA
– are spheres of lipid molecules surrounding an aqueous interior (bilayer); limited amount of DNA will fit in it
liposome
– are lipids with positive charge so attract both DNA and cell wall
No immune response, but not as efficient as virus at transferring DNA
lipolplex
Direct injection of – into muscle cells was found to be expressed
Could be used to make insulin, blood clotting factors or as vaccines to viruses.
naked DNA
Ex vivo usually targets – which are stem cells that generate all other blood cells
hematopoietic stem cells (HSC)
X-linked severe combined immunodeficiency (SCID-X1)
β-thalessemia
WAS – an immune difficiency disorder
are associated with
HSC
is an RNA-mediated mechanism for regulating gene expression in eukaryotes (regularly used to silence specific genes)
RNA interference
RNAi is a cellular defense against viruses and –
transposable elements
viral RNA uses – to become dsRNA virus
RNA dependent RNA polymerase
– cuts dsRNA into 21-25 bp fragments
Dicer
siRNA fragments created by Dicer bind to – which denatures one of the two strands
RISC
guide RNA remains bound to RISC and then the – slices other RNA that is complementary to the guide strand
argonaute
RNAi is used in research to test hypothesis of gene function this is known as –
knock-down
RNAi is used in therapeutics to – RNA of overexpressed genes
degrade
RNA is used in therapeutics to target specific –
viruses (HIV)
T/F: RNAi destroys genome
false
RNAi methods: – of siRNAs directly into cells
transfection
RNAi method: introduction of – synthesizes RNA identical to endogenous gene of interest (siRNAs made in organism)
vectors
T/F: small interfering RNA (siRNA) come from exogenous sources or from other endogenous transcription
true
Abnormal over-expression of – lead to diseases such as age-related macular degeneration (AMD)
VEGF (vascular endothelial growth factor)
VEGF is a protein required for blood vessel formation
Excessive blood vessels damage retina
– VEGF should solve the problem
silencing
problems associated with RNAi include the – of sRNAs and adequate delivery systems
stability
In RNAi silencing, specificity is provided by –
siRNA
go in and make changes at a particular site, generally not replacing large amounts
genome editing
ancient bacterial immune system which identifies the DNA of invading viruses and sends in an endonuclease, like Cas9, to shew it up
CRISPRS
– can be reprogrammed to recognize nearly any sequence of DNA
just need the right RNA sequence to search a strand of DNA for where to cut
Cas9
The – regions are homologous to viruses
spacer
The – are important for binding the Cascade complex
repeats
Spacer RNA directs complex to – DNA or RNA
homologous
In E.coli system, cascade complex has – activity
Dnase
In Pyrococcus furiosus system, Cmr-complex has – activity
Rnase
One can now use the CRISPR system in eukaryotes to do gene knockouts, gene replacements, and –
gene “drive”
In crop plants, the genome can be edited and then the – can be segregated away, leaving the plant transgene free
Cas9 plasmid
selfish genes that appear more frequently in offspring than normal genes, which have about a 50-50 chance of being passed on
gene drives
T/F: A gene drive always ends up in all offspring, even if only one parent has it;
if given enough generations, it will eventually spread through the entire population
true
Gene drive: Fight Malaria by changing the mosquitos that are part of their life cycle.
One group modified females to become –
sterile
Gene drive: Fight Malaria by changing the mosquitos that are part of their life cycle. The other group modified mosquitos by inserting two – directed against the malarial parasite.
antibody genes
Transgenic to fight Zika virus larvae died due to high expression of –
lethal transgene
