forward genetics: analyzing and determining the mutation causing a certain phenotype
reverse genetics: causing a mutation and analyzing the resulting phenotype
gene knockout: the targeted removal/inactivation of a specific gene
starting/maintaining a cell line: isolating individual cells from an individual by disrupting the
extracellular matrix and cell-junctions with proteolytic enzymes, then the cells are cultured
and maintained on a solid and coated surface that they can adhere to
fluorescence-activated cell sorter: fluorescence molecules are coupled to antibodies to make
specific cells fluorescent, single drops (containing 1 cell) are given a charge, based on if they
are fluorescent or not, the the cells are sorted into collection tubes by deflection, this is done
with an electric field
laser micro-dissection microscope: a specific tissue of interest is ‘cut out’ by a laser, then the
tissue is catapulted into a clean container with use of a different laser beam
cell line: a cell line is obtained from repeated culturing (growth vectors used) of isolated
individual cells
ca2+ phosphate co-precipitation: upon addition of calcium and phosphate, DNA precipitates,
these precipitates are added to a cell line and integrate in the cells and then in the nucleus of
these cells
electroporation: the plasmid (goi and marker gene) and cells are mixed and then a voltage is
applied, because of this voltage, the cells become semi-permeable and the DNA can enter
the cells and its nucleus and therefore integrate into the genome of the cell, to yield stable
expression a selection is performed
lipofection: a construct is mixed with lipid molecules upon which liposomes containing the
DNA are formed, these liposomes are added to the cells and adhere and fuse with its
membrane, therefore the DNA can enter the cell and its nucleus
viral vector adenovirus: dsDNA, linear episomes, minimal integration, cloning capacity,
replication and pathogenicity
viral vector retrovirus: inserts DNA copy into DNA of the host cell to change the genome of
the cell, applied in gene therapy
packaging cell line: when plasmids containing packaging genes are added to a cell line, and
integrate successfully in the genome of the cells, the packaging genes are expressed and
you end up with a cell line expressing packaging proteins, this cell line is used to generate
recombinant AAV virus particles
viral vector adeno-associated virus: doesn’t integrate in genome, replication defective
contains 3 ORFs (rep, cap, AAP), gene of interest is inserted between ITRs (- rep and cap)
and that yields a recombinant AAV
micro-injection in fertilized egg: DNA is inserted into the pronucleus of a fertilized egg via
injection with a microneedle
gene therapy: treatment of a hereditary disease by inserting a transgenic construct to
replace the endogenous gene causing the disease
direct delivery (AAV and non-viral vector): happens in vivo with vector injected into patient
stem cell-based delivery: stem cells are isolated from the organism and modified, placing
these back into the organism results a non-chimeric individual
embryonic stem cells: embryonic stem cells can convert to all tissue types but cannot give
rise to an entire organism
chimerism: chimerism is obtained when an organism is composed of cells with two or more
different genotypes
induced pluripotent stem cells: a retrovirus, containing 4 proteins, infects a somatic cell and
these proteins lead to the conversion of the somatic cell into a pluripotent stem cell
reverse genetics: causing a mutation and analyzing the resulting phenotype
gene knockout: the targeted removal/inactivation of a specific gene
starting/maintaining a cell line: isolating individual cells from an individual by disrupting the
extracellular matrix and cell-junctions with proteolytic enzymes, then the cells are cultured
and maintained on a solid and coated surface that they can adhere to
fluorescence-activated cell sorter: fluorescence molecules are coupled to antibodies to make
specific cells fluorescent, single drops (containing 1 cell) are given a charge, based on if they
are fluorescent or not, the the cells are sorted into collection tubes by deflection, this is done
with an electric field
laser micro-dissection microscope: a specific tissue of interest is ‘cut out’ by a laser, then the
tissue is catapulted into a clean container with use of a different laser beam
cell line: a cell line is obtained from repeated culturing (growth vectors used) of isolated
individual cells
ca2+ phosphate co-precipitation: upon addition of calcium and phosphate, DNA precipitates,
these precipitates are added to a cell line and integrate in the cells and then in the nucleus of
these cells
electroporation: the plasmid (goi and marker gene) and cells are mixed and then a voltage is
applied, because of this voltage, the cells become semi-permeable and the DNA can enter
the cells and its nucleus and therefore integrate into the genome of the cell, to yield stable
expression a selection is performed
lipofection: a construct is mixed with lipid molecules upon which liposomes containing the
DNA are formed, these liposomes are added to the cells and adhere and fuse with its
membrane, therefore the DNA can enter the cell and its nucleus
viral vector adenovirus: dsDNA, linear episomes, minimal integration, cloning capacity,
replication and pathogenicity
viral vector retrovirus: inserts DNA copy into DNA of the host cell to change the genome of
the cell, applied in gene therapy
packaging cell line: when plasmids containing packaging genes are added to a cell line, and
integrate successfully in the genome of the cells, the packaging genes are expressed and
you end up with a cell line expressing packaging proteins, this cell line is used to generate
recombinant AAV virus particles
viral vector adeno-associated virus: doesn’t integrate in genome, replication defective
contains 3 ORFs (rep, cap, AAP), gene of interest is inserted between ITRs (- rep and cap)
and that yields a recombinant AAV
micro-injection in fertilized egg: DNA is inserted into the pronucleus of a fertilized egg via
injection with a microneedle
gene therapy: treatment of a hereditary disease by inserting a transgenic construct to
replace the endogenous gene causing the disease
direct delivery (AAV and non-viral vector): happens in vivo with vector injected into patient
stem cell-based delivery: stem cells are isolated from the organism and modified, placing
these back into the organism results a non-chimeric individual
embryonic stem cells: embryonic stem cells can convert to all tissue types but cannot give
rise to an entire organism
chimerism: chimerism is obtained when an organism is composed of cells with two or more
different genotypes
induced pluripotent stem cells: a retrovirus, containing 4 proteins, infects a somatic cell and
these proteins lead to the conversion of the somatic cell into a pluripotent stem cell
