Bi6721c Special Methods of Microorganisms Analysis I. - practical course

Faculty of Science
Spring 2011 - only for the accreditation
Extent and Intensity
0/3/0. 3 credit(s). Type of Completion: z (credit).
Teacher(s)
doc. RNDr. Alena Španová, CSc. (seminar tutor)
doc. Ing. Bohuslav Rittich, CSc. (seminar tutor)
RNDr. Aleš Kovařík, CSc. (seminar tutor)
Guaranteed by
doc. Ing. Bohuslav Rittich, CSc.
Department of Experimental Biology – Biology Section – Faculty of Science
Contact Person: doc. Ing. Bohuslav Rittich, CSc.
Prerequisites
NOW( Bi6721 Spec. Meth. Microorg. Anal. I. )
The course is destined for students interested in practical knowledge of method PCR (polymerase chain reaction).
Course Enrolment Limitations
The course is also offered to the students of the fields other than those the course is directly associated with.
fields of study / plans the course is directly associated with
Course objectives
At the end of this course, the students should be able practically use the method of polymerase chain reaction for the identification of bacteria.
Syllabus
  • 1. Safety of work in microbiology laboratory. Pipeting of small volumes of solvents using pippetmans. Preparation of media for cultivation of bacteria.
  • 2. Preparation of Salmonella Typhimurium LB5000 cells for isolation of bacterial DNA. Cell lysis.
  • 3. DNA deproteination using phenol extraction. Precipitation of DNA with ethanol.
  • 4. Estimation of DNA integrity using agarose gel electrophoresis of isolated DNA. DNA visualisation and gel documentation.
  • 5. Estimation of DNA concentration and purity. Preparation of DNA in concentration suitable for PCR.
  • 6. The work flow in PCR laboratory. Preparation of PCR mixture for specific amplification of gene Salmonella using purified DNA as DNA matrix. PCR reaction.
  • 7. Detection of PCR product using agarose gel electrophoresis.
  • 8. Estimation of amplicon length using known standards.
  • 9. Preparation of PCR mixture from trude cell lysates. Positive and negative controls. PCR and detection of PCR produkt.
  • 10.Imunomagnetic separation (IMS) of cells Salmonella. IMS-cultivation (IMS-CM) of cells Salmonella. IMS-polymerase chain reaction (IMS-PCR) with cells Salmonella. Estimation of PCR sensitivity.
  • 11.I PCR products detection using agarose gel electropjoresis.
  • 12. and 13. Identification of Salmonella cells in unknown sample using PCR.
  • 14. Evaluation of protocols.Test.
Literature
  • F.Sambrook and D.W. Russell Molecular Cloning. A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press. 2001
Teaching methods
Each student works independently and processes his or her own sample during the laboratory course. The results of their experiments must be evaluated in a protocol. The protocols (4 in number) are elaborated in the form of a poster according to the following scheme: Introduction, Aim of work, Material and methods, Results, Discussion, Conclusion. In the protocol each student describes, compares, analyses, evaluates, and discusses his or her own results.
Assessment methods
The course is closed by a course-unit credit. The students obtain their credits for active attendance at the course and for the preparation of protocols (4). In the course of the credit exercise, each student has to identify target bacteria in an unknown sample using the PCR method.
Language of instruction
Czech
Follow-Up Courses
Further comments (probably available only in Czech)
The course is taught annually.
The course is taught: every week.
Information on course enrolment limitations: Přednost mají posluchači specializace mikrobiologie.
The course is also listed under the following terms Spring 2008 - for the purpose of the accreditation, Spring 2003, Spring 2004, Spring 2005, Spring 2006, Spring 2007, Spring 2008, Spring 2009, Spring 2010, Spring 2011, spring 2012 - acreditation, Spring 2014, Spring 2016, Spring 2017, spring 2018, Spring 2019, Spring 2020, Spring 2021, Spring 2022, Spring 2024, Spring 2025.