Bi7722c Methods of Microorganisms Analysis II - practical course

Faculty of Science
Autumn 2002
Extent and Intensity
0/4/0. 4 credit(s). Type of Completion: z (credit).
Teacher(s)
doc. Ing. Bohuslav Rittich, CSc. (seminar tutor)
doc. RNDr. Alena Španová, CSc. (seminar tutor)
Guaranteed by
doc. RNDr. Alena Španová, CSc.
Department of Experimental Biology – Biology Section – Faculty of Science
Contact Person: doc. RNDr. Alena Španová, CSc.
Prerequisites (in Czech)
Bi6721 Spec. Meth. Microorg. Anal. I.
Course Enrolment Limitations
The course is also offered to the students of the fields other than those the course is directly associated with.
The capacity limit for the course is 12 student(s).
Current registration and enrolment status: enrolled: 0/12, only registered: 0/12, only registered with preference (fields directly associated with the programme): 0/12
fields of study / plans the course is directly associated with
Course objectives
I. Work with recombinant DNA.Cultivation of bacterial cells E.coli JM109(pUC19::dim1), preparation of solutions for plasmid DNA isolation.Estimation of DNA quantity using gel electrophoresis with suitable standards. Change of buffer in which DNA is solved. Transfer of recombinant and nonrecombinant plasmid DNAs into bacterial cells E.coli JM109 using electrotransformation. Selection of electrotransformants carrying recombinant and nonrecombinant plasmid DNAs. Evaluation of electrotransformation. II. DNA/DNA hybridisation. Preparation of DNA probe from plasmid DNA pUC19:dim1 - nonradioactive labelling with digoxigenine. Preparation and carrying out of dot blot and Southern blot. Carrying out of DNA/DNA hybridisation at high stringency. Immunological detection of hybridisation product. Evaluation of hybridisation.
Syllabus
  • 1. Safety of work in microbiological and molecular biotechnical laboratory. 2. Preparation of materials for isolation of recombinant plasmid DNA.Inoculation of bacterial cells, preparation of solutions. 3. Isolation of recombinant DNA pUC19::dim1 from E. coli JM109(pUC19::dim1) 4. Change of buffer in which DNA is solved.Agarose gel electrophoresis of isolated DNA. 5. Transfer of recombinant and non recombinant plasmid DNAs into bacterial cells E. coli JM109 using electrotransformation. 6.Plating and selection of electrotransformants carrying recombinant and nonrecombinant plasmid DNA. 7. Evaluation of electrotransformation. Preparation of southern blot. 8. Preparation of DNA probe from plasmid DNA with neradioactive labelling with digoxigenine. 9. Preparation and carrying out of dot blot and Southern blot. 10. Gel electrophoresis of recombinant plasmid DNA with labelled and nonlabelled controls. 11. Southern blot. 12. DNA/DNA hybridisation at high stringent conditions. 13. Immunological detection of hybridisation products. 14. Evaluation of hybridisation. 15. Test. Control of protocols.
Literature
  • F. Sambrook, R.W. Russell Molecular Cloning. A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press. 2001.
Language of instruction
Czech
Further Comments
The course is taught annually.
The course is taught: every week.
  • Enrolment Statistics (recent)
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