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GENETIC TECHNOLOGY
1) GENETIC RECOMBINATION
1.
2.
3.
4.
Remove bacterial plasmid with
restriction enzymes
Add in gene of interest (plasmid
is now recombinant DNA
molecule)
Put back into bacteria
Many reproductive cycles later
= amplification of gene &
protein it makes
RESTRICTION ENZYMES
 Cut
up foreign DNA
 Very specific
 Recognize short nucleotide
sequences (restriction site)  cut
at specific points within sequence
 Same sequence found on both
strands, running antiparallel
RESTRICTION ENZYMES
 Enzyme
cuts phosphodiester
bonds of strands
 These restriction fragments are
double stranded with single
stranded ends (“sticky ends”)
 Bacteria’s own DNA is methylated
to protect itself
RESTRICTION ENZYMES
 Single
strands will
hydrogen bond with
other complementary
“sticky ends”
 Bonds made permanent
with DNA ligase
 Now we have
recombinant DNA
FROM RESTRICTION ENZYMES
TO PLASMID MAPS
 PM
show how different REs act
upon a plasmid
 Pictorial representation of the
different lengths
of pieces
remaining
after the
REs worked
FROM RESTRICTION ENZYMES
TO PLASMID MAPS
 Procedure
 Think
of plasmid as
clock – from 12 to 12 =
total # base pairs
 Approximate location
of cut based on base pair
fragment length
 Use logic to solve
 Double check based on data
2) DNA ANALYSIS GEL ELECTROPHORESIS
 Sequence
of entire genome 
genomics
 Begins with gel electrophoresis
 Sorts DNA based on size &
charge
 Can combine
with specific
probes to
label
particular
DNA
bands
3) POLYMERASE CHAIN REACTION
(PCR)
 Can
quickly amplify specific DNA
without using cells
 DNA of interest incubated with
DNA polymerase, nucleotides, &
ss primer DNA for synthesis
 DNA heated  strands separate
 Cool  primers bond
 DNA polymerase adds to 3’ end
of each primer
 Repeat
4) GENOME ANALYSIS
 DNA
microarray:
ssDNA fragments
fixed to slide that
are then labeled
with fluorescent
cDNA
 Compare genes of
species  attempt to
uncover gene function
5) GENOME ANALYSIS - GENE
FUNCTION
 To
determine, turn gene off – see
what happens
 To turn off: RNA interference
(RNAi)
 Synthetic ds RNA matches gene
sequence – binds to mRNA
 Triggers breakdown of mRNA 
no protein made
 Remove to turn on again
THE FUTURE
 Proteomics:
study of full protein
sets
 Study of variations among the
species
 Form of single nucleotide
polymorphisms (SNPs)
Single base-pair variations
One per 1000 bp
DNA TECHNOLOGY
APPLICATIONS
 Disease
Diagnosis
 Use PCR & labeled nucleic acid
probes to detect pathogens (ex:
HIV)
 Identification of harmful
alleles before birth
DNA TECHNOLOGY
APPLICATIONS
 Human
Gene Therapy
 Alteration of genes
 Replace defective gene with
normal one  put into cells
that keep dividing
Appears to be temporary
 Raises ethical questions
DNA TECHNOLOGY
APPLICATIONS
 Pharmaceutical
Products
 Use vector DNA to create
human insulin, HGH, TPA, etc
 Recombinant DNA to make
vaccine without using actual
pathogen
DNA TECHNOLOGY
APPLICATIONS
 Forensics
 Microsatellite
DNA highly
variable between individuals
Called simple tandem repeats
(STR)
 Environmental
 Genetically engineered microbes
to degrade toxic waste
DNA TECHNOLOGY
APPLICATIONS
 Agriculture
 Transgenic
organisms: carry
genes from another species
Makes “super” species
Remove egg  fertilize in vitro
 inject desired DNA into egg
nuclei  cell will grow &
express gene  egg put into
surrogate
DNA TECHNOLOGY
APPLICATIONS
 Plants
Vector is recombinant Ti
plasmid – inserts into plant
genome  cell grows into
complete plant
Can increase
nutritional
value
