BLIRT EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique

Aim: to introduce students to the PCR-RFLP method (Restriction Fragment Length Polymorphism).

EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique
EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique
EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique
Name Part Number Pack Size
EasyGenotyping PCR-RFLP S. aureus - genotyping using PCR-RFLP technique DY87 1 exercise
  DY875 5 exercises

 

EasyGenotyping PCR-RFLP S. aureus - educational kit for bacteria strains genotyping using PCR-RFLP (PCR Restriction Fragment Length Polymorphism) technique.

 Aim: to introduce students to the PCR-RFLP method (Restriction Fragment Length Polymorphism). It is used for intra- and interspecies genotyping of bacterial and fungal strains. The method is carried out in 2 steps. In the first step a region of interest is amplified and a PCR product of specified size is obtained. In the next step it is digested with one or more restriction enzymes. As a result of the digestion, a restriction profile is obtained, specific for a particular isolate, which allows to assign the strain to a particular specie or genotype. The aim of this class is to conduct genotyping of the supplied genomic DNA of Staphylococcus aureus isolates and to assign them to the individual genotype groups.

The kit is designed to perform experiments during three laboratory lessons (I - PCR set-up - 1.5-2 hours; II - electrophoresis and restriction digestion - 2 hours; III - electrophoresis - 2 hours). We suggest to perform laboratory experiments in six groups of two students.

EasyGenotyping PCR-RFLP S. aureus kit components (1 exercise, DY87)
A) 6 sets of PCR reagents:
- 6 tubes with PCR buffer (each for 10 reactions)
- 6 tubes with MgCl2 solution (each for 10 reactions)
- 6 tubes with ready-to-use premixed dNTPs solution (each for 10 reactions)
- 6 tubes with forward primer solution (each for 10 reactions)
- 6 tubes with reverse primer solution (each for 10 reactions)
- 6 tubes with PCR grade water
- 6 tubes with thermostable polymerase (each for 10 reactions)
- 12 tubes with genomic DNA of 12 Staphylococcus aureus isolates for analysis (each for 3 reactions)
B) 6 sets of reagents for digestion reaction:
- 6 tubes with digestion reaction mix (each for 12 reactions)
- 1 tube with restriction enzyme (for 12 reactions)
C) Moreover:

- 7 laboratory protocols (one with additional information for a teacher)

- Agarose (5 g)

- 50x TAE buffer for agarose gel electrophoresis (50 ml)

- Ready-to-use M100-1000 DNA Ladder for 10 electrophoretic lanes

- Ready-to-use M100-500 DNA Ladder for 10 electrophoretic lanes

- DNA Gel Loading Buffer - 6x GREEN (0.5 ml)

EasyGenotyping PCR-RFLP S. aureus kit components (5 exercises, DY875):
A) 6 sets of PCR reagents:
- 6 tubes with PCR buffer (each for 50 reactions)
- 6 tubes with MgCl2 solution (each for 50 reactions)
- 6 tubes with ready-to-use premixed dNTPs solution (each for 50 reactions)
- 6 tubes with forward primer solution (each for 50 reactions)
- 6 tubes with reverse primer solution (each for 50 reactions)
- 6 tubes with PCR grade water
- 6 tubes with thermostable polymerase (each for 50 reactions)
- 12 tubes with genomic DNA of 12 Staphylococcus aureus isolates for analysis (each for 15 reactions)
B) 6 sets of reagents for digestion reaction:
- 6 tubes with digestion reaction mix (each for 60 reactions)
- 1 tube with restriction enzyme (for 60 reactions)
C) Moreover:

- 7 laboratory protocols (one with additional information for a teacher)

- Agarose (25 g)

- 50x TAE buffer for agarose gel electrophoresis (250 ml)

- Ready-to-use M100-1000 DNA Ladder for 50 electrophoretic lanes

- Ready-to-use M100-500 DNA Ladder for 50 electrophoretic lanes

- DNA Gel Loading Buffer - 6x GREEN (1 ml)

Additional equipment and materials required (not included in the kit):

- Automatic pipettes with appropriate tips

- PCR and 1.5 ml Eppendorf tubes

- UV transilluminator or lamp

- Thermal cycler

- Microwave oven or laboratory burner

- Agarose electrophoresis equipment (apparatus, power supply)

- Ethidium bromide solution or other stain for DNA visualization

- Nitrile gloves

Shipping condition

Shipping on dry or blue ice.