lecture 37 - sae 5 Flashcards
describe somatic cell fusion
gene mapping
describe gene therapy
conventional gene therapy
genome editing and ips
describe cell hybridization/fusion
whole cell fusion
virus binds 2 cells – viral membrane fuses membranes of cells – makes bridge then fused together = now hybrid
high confluence - touch each other
describe hybrids created
hybrids containing genomes from the 2 diff cell lines
genomes of fused cells are unstable = randomly keep or lose chromosomes
hybrid between human and rodent = rodent chromosomes mostly kept and most human chromosomes lost = used to map gene of interest in human
is cell fusion efficient
noooo highly inefficient
selectable marker needed to identify hybrids
describe hat medium
hypoxanthine aminopterin thymidine
natural marker selection
describe de novo pathway
blocked by aminopterin
noble pathway
major dna precursors
makes newly synthesized dna
describe salvage pathway
minor dna precursors
salvage pathway like recycles
can live without as long as major INTACT
requires hprt for dgtp and tk for dttp = 2 genes become essential when major not working
if can provide these precursors then cell will live - even tho major pathway gone
define hypoxanthine and thymidine
hypoxanthine = converted to guanine by hypoxanthine guanine phosphoribosyltransferase
thymidine = phoshorylated by thymidine kinase
describe fusion of the hprt and tk lines
fusion of tk deficient and hprt deficient cell = new fusion of 2 cell lines = kinda like complementation test - so now cell has tk and hprt
now full set of enzymes = survive off of minor pathway
describe mapping measles virus receptor in humans - experiment
measles can only bind to primates not rodents
make hybrids between primate and rodent - some human chroms kept but most lost –> identify cells infected by measles –> determine which human chroms kept in those cells (easy since human chrom structure diff - morphologically diff from rodents)
if receptors for measles lost then no measles
describe mapping measles virus receptor in humans - results
look at all human chroms
3/9 can be affected by measles = look at common number so chrom 1 is common in all that have measles
Probably where receptor for measles encoded
what is gene therapy
involves adding normal wild type copy of a gene to genome of an individual carrying defective mutated copies of the gene - often recessive mutations
finding defective genes
only few of 4000 inherited human diseases are treatable
name the 2 types of viral vectors
derived from adenovirus
derived from retrovirus
describe viral vector derived from adenovirus
will infect all cells even non dividing ones
vectors will not be integrated in genome = transgene diluted and eventually lost = not permanent but faster
describe viral vector derived from retrovirus
many will infect only diving cells but others (hiv) can infect without host cell division
transgene and viral vector will be incorporated into genome of infected cells - more stable
describe somatic gene editing
limited to tissue - targets genes in specific types of cells
edited gene is only contained in target cell type
any changes limited to treated individual - edited gene not passed down to future generations
describe germline gene editing
made early in development so any changes copies into all of new cells = alters genome
copied in every cell = includes sperm and egg
will be passed on to future gens
new - mostly not approved since dont know effects
which type of gene editing is more common
somatic gene editing
what is somatic cell gene therapy
transfer of a gene in somatic cells
what is germline gene therapy
transfer of a gene in all cells of an organism through germline transmission
describe in vivo vs ex vivo gene therapy
in vivo = inject and hope it hits right cells
ex vivo = isolate hematopoietic target cells and alter them and then put them back in person = one can better control and monitor efficiency of gene transfer
what does ada-scid stand for
adenosine deaminase - severe combined immunodeficiency disease
rare autosomal disease of immune system - bubble boy disease, affected individuals have essentially no immune system
describe ada-scid
in absence of ada deoxyadenosine accumulates in t lymphocytes and eventually kills these cells - t lymphocytes responsible for stimulation of antibody producing b cells
first disease treated with gene therapy - 1990
most common treatment = bone marrow transplant
