Professional Documents
Culture Documents
of Medicine
Department of Medical Biology Universitas Swadaya Gunung Jati
2017
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alleles at a G g G G g g
gene locus
Replication
R r R R r r
S s S S s s
t T t t T T
Homologous chromosomes
3
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4
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Genetic engineering
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§ Biotechnology
Technic which applies biological system (in organisms, cellular and molecular level) aiming
to obtain the expected result according to human needs.
Biomedical importance
1. Rational approach to understanding the molecular 4. Human proteins can be produced
basis of a number of diseases in abundance for therapy
E.g: muscular dystrophy, cystic fibrosis, cancer, diabetes E.g: Insulin, growth hormone
to pharmacological therapeutics
E.g: Ebola test, familial cancer 6. Forensic medicine
E.g: Method for identifying criminal
3. Gene therapy for potentially curing diseases suspect
Human
genome
1. Gene identification
§ Polymerase chain reaction (PCR) & electrophoresis
§ Restriction fragment length polymorphism (RFLP)
§ Hybridization: Western, southern, northern blot
§ DNA sequencing
2. Gene cloning
3. Transgenic animal
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Target DNA
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Step 1 (94oC)
Denaturation
dsDNA to ssDNA
Step 2 (56-60oC)
Annealing
Primers onto template
Step 3 (72oC)
Extension
dNTPs extend the 2nd strand
Process
Denature
Anneal Primer
Replicate
DNA
§ Requires:
§ DNA template
§ A pair of primers
§ DNA polymerase (thermostable; obtained from
thermophilic bacteria) à Thermus aquaticus
§ Taq polymerase
§ dNTP
§ Thermal cycler (PCR machine)
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Gene identification:
§ To separate charged cells/molecules component Electrophoresis
(protein, nucleotide).
§ Principle: charged molecule in electric field will
migrate to opposite electrode. DNA is negatively
charged, will migrate to anode.
§ Samples are loaded into well in one of the plate
site
§ Velocity of the migration depends on:
§ Charged level
large moderate small
§ Molecule’s size
§ Molecule’s shape
§ Supporting media: Agarose/polyacrylamide gel &
Buffer
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Electrophoresis
N O N O N O N O M
1 2 3 4 5 6 7 8 9
800 bp 1000 bp
472 bp
500 bp
326 bp
311 bp
§ E.g:
Polymorphism in exon 10 of FSH receptor gene in position 680th
nucleotide à resulting in 2 different amino acids: Ser & Asn.
AACC CGGGAG AACC AGGGAG
TTGGGC CCTC TTGGGT CCTC
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1 kb a. Marker
b. Ser/Ser
500bp c. Asn/Asn
d. Ser/Asn
100bp
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Southern blot
hybridization
Hybridization
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Blunt ends
Sticky ends
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Restriction enzymes
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DNA polymerase I
§ Isolated from bacteria
§ Incorporates nucleotides to complementary template DNA to synthesizing
new dsDNA; generates blunt ends from sticky ends.
Murray RK, et al. 2009. A LANGE medical book: Harper’s illustrated biochemistry. 28th ed. New York: The McGraw-Hill Companies, Inc
“Tempora Mutantur Nos et Mutantur In Illis”
“Time changes and we change with them”