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biology
modern genetics
7.1 using gene sequencing
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abdul ahmed
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Cards (13)
genome
?
complete set of
genetic
information and
material
in the
cells
of an
organism.
what is dna sequencing?
identifying
base
sequences of
nucleotide
in dna fragment
Dna
samples are amplified by
polymerase chain reactions
(PCR)
what is in the reaction mixture of PCR?
target
DNA
(dna to be amplified)
free
nucleotides
dna
polymerase
(catalyse formation of
phosphodiester
bonds)
primers
what are primers?
short
nucleotide
base
sequence -
complimentary
to the dna that is going to be
amplified.
allows
dna polymerase
to
anneal
to template strand and begin
replication
Steps
of PCR
DNA strand
heated
to
95
degrees
separates
DNA strands and
hydrogen
bonds break
cooled
to
55
degrees
allows
primers
to
anneal
to DNA strand
heated
to
70
degrees
to activate
DNA polymerase
and allow nucleotides to
join.
Steps of Gel electrophoresis
Dna
sample is
amplified
by
PCR
Restriction
enzyme breaks DNA into
fragments
Stained with
fluorescent
dye
Potential
difference used
Dna shows up as
bands
No of bands links to
similarity
steps of southern blotting
DNA sample is
amplified
by
PCR
Restriction
enzyme breaks
DNA
into
fragments
gel placed in an
alkali
solution to breaks
hydrogen
bonds between DNA
bases
so strands separate.
thin nylon sheet placed on gel
placed in bag with probes
probes anneal to DNA with complimentary base
looked at through x-ray
what are
probes
?
short
single
stranded
DNA
labelled with
radioactive phosphorus
locates
bands
on nylon
locates specific
alleles
and
base sequences
uses of amplified DNA samples
DNA
sequencing
(predicts amino acid sequence/ genetic disease)
DNA
profiling
(forensics)
steps
of DNA sequencing (termination):
4
separate reactions containing...
template DNA, DNA
polymerase
, primers,
4
nucleotides
PCR occurs in
each
reaction tube
terminator
nucleotide added +
replication
stops
DNA fragments separated by size using
gel
electropheresis
viewed under
UV
strand moved furthest is first base in
complimentary
strand
what
is satellite DNA?
in non coding regions
long strands of DNA containing
short tandem repeats
(STR)
Steps
of DNA Profiling:
DNA sample collected and amplified by
PCR
satellite DNA cut by
restriction
enzymes to form fragments
fragment
length
depends on length of
short
tandem repeats
fragment separated using
gel electrophoresis
southern
blotting used to identify target sequence.