Course detail

Laboratory Classes in Molecular Biotechnology

FCH-MC_MOB_PAcad. year: 2023/2024

The course is focused on practical mastering of method polymerase chain reaction in identification of bacteria of genus Lactobacillus in milk product (probiotic drink). PCR-ready DNA is isolated from the product using magnetic microspheres. PCR mixtures are prepared after estimation of DNA concentration and quality. DNA is amplified in conventional PCR and in real-time PCR. Working out of for protocols in the form of posters is part of the course.

Language of instruction

Czech

Number of ECTS credits

Mode of study

Not applicable.

Guarantor

prof. Mgr. Václav Brázda, Ph.D.

Department

Institute of Food Science and Biotechnology (ÚCHPBT)

Entry knowledge

It is recomended to pass out the lecture Bioanalytical methods.

Rules for evaluation and completion of the course

Laboratory course is finished by test and cotrol of protocols.
Attendance in laboratory course is obligatory.

Aims

The aim of the laboratory course is to cover methodologies used in the work with DNA particularly its amplification in PCR. The next aim covers the preparation of poster.
Practical knowledges of some basis experimental techniques used in DNA research.

Study aids

Not applicable.

Prerequisites and corequisites

Not applicable.

Basic literature

Španová A., Rittich B.: Analýza vybraných druhů baktérií mléčného kvašení pomocí metod molekulární biologie (CS)

Type of course unit

Guided consultation in combined form of studies

52 hod., optionally

Teacher / Lecturer

prof. Mgr. Václav Brázda, Ph.D.

Laboratory exercise

52 hod., compulsory

Teacher / Lecturer

Ing. Lenka Fialová, Ph.D.
Ing. Lucie Šislerová
Ing. Libuše Kratochvilová
Ing. Michaela Dobrovolná

Syllabus

1. Safety of the work in laboratory. Arrangement of PCR laboratory. Pipetting of small volumes.
2. Cultivation of Lactobacillus cells for isolation of bacterial DNA. Estimation of purity of bacterial culture.
Cell lysis and preparing of crude cell lysate. Removing of proteins using phenol extraction. Precipitation of DNA with ethanol. Determination of DNA integrity using agarose gel electrophoresis. Preparation of a protocol in the form of poster.
3. Spectrophotometric estimation of DNA concentration and purity. Preparation of PCR mixture for amplification of specific part of Lactobacillus DNA using purified DNA. Preparation of negative control. PCR amplification. Detection of PCR product using agarose gel electrophoresis. Estimation of length of amplicon with help of DNA standard and programme Bionumerics. Preparation of a protocol in the form of poster.
4. Isolation of whole DNA from crude cell lysates of dairy product (probiotic drink) using magnetic particles P(HEMA-co-GMA). Amplification of DNA by PCR in real time. Preparation of a protocol in the form of poster.
5. Introduction to bioinformatics - lecture a practical excersize on PC.
6. Test and control of protocols.

Elearning

eLearning: currently opened course