If the organism is:
Difficult/costly to culture
Present in low concentration in clinical
Hazardous to propagate in the lab
It is amplification of a specific DNA
sequence using specific primers.
3. PCR is:
Sensitive (1 copy → million copies)
Specific (specific primers)
Rapid, efficient, reproducible
4. Primers are:
2- Single stranded
3- Short sequence
4- Complementary to the flanks of desired
region of target DNA
5. PCR Technique
Extracted N.A. + 3 items:
1- Two primers (forward & reverse)
2- Taq polymerase
3- Nucleotide pool
1- Denaturation (95°C)
2- Primer annealing (40-60°C)
3- Primer extension (65-75oC) .
6. Thermal Cyclers
7. 3 methods for detection of amplified
1- Gel electrophoresis.
8. Gel electrophoresis:
• DNA fragments migrate in the gel
• Migration rate depends on size of DNA fragments
(length), smaller fragments → more rapid
• A dye (in the gel) binds to DNA fragments →
allows visualization by UV light
9. Gel electrophoresis
10. Gel electrophoresis
11. In vitro
Antibiotic Susceptibility Tests
12. Antibiotic Susceptibility Tests
Disc Diffusion Method: for routine use.
Test org. on surface of media
Zones of Inhibition
Susceptibility to antibiotic
13. Disc Diffusion Method
5 ,3 ,2 ,1
1-All of the following are characters of primers
b. Double stranded
c. Short sequence
d. Complementary to the flanks of target DNA
e. Used in PCR
15. 2- The amplified DNA products may be detected by
any of the following EXCEPT:
b.Disc diffusion method
16. 3- Regarding the PCR technique all the following
are included in the reaction EXCEPT:
a.Two primers (forward & reverse).
c.Extracted N. A. (target).
17. 4- Formation of clear zones around antibiotic discs is
a.Resistance of the bacterium
b.Short incubation period
c.The expiry of the antibiotic discs
d.Inhibition of bacterial growth around the discs
e.Low concentration of the antibiotic
18. 5- Regarding gel electrophoresis:
a.It is a method of amplification of DNA.
b.It is a method of testing electrical forces of
c.Visualization of fragments is done by infrared light.
d.Smaller fragments migrate faster.
e.DNA visualization does not need a dye.