Bi8313 Genetic Engineering - Laboratory Course

Faculty of Science
Spring 2023
Extent and Intensity
0/2/0. 2 credit(s). Type of Completion: z (credit).
Teacher(s)
Mgr. Ivana Mašlaňová, Ph.D. (seminar tutor)
prof. RNDr. Roman Pantůček, Ph.D. (seminar tutor)
Mgr. Lucie Kuntová, Ph.D. (seminar tutor)
Mgr. Jiří Holoubek (assistant)
Mgr. Radka Obořilová (assistant)
Mgr. Hana Šimečková (assistant)
Guaranteed by
prof. RNDr. Roman Pantůček, Ph.D.
Department of Experimental Biology - Biology Section - Faculty of Science
Contact Person: prof. RNDr. Roman Pantůček, Ph.D.
Supplier department: Department of Experimental Biology - Biology Section - Faculty of Science
Prerequisites
! B6492 Molecular biology - practice 2 && ( Ex_2979 Metody molekulární biologie || Ex_2980 Metody molekulární genetiky || B6390 Molecular Methods || B6400 Methods of molecular biology || Bi6400 Methods of molecular biology ) && ( Bi6405 Molecular Methods - practice || B6405 Molecular Methods - practice || Bi6400c Molecular Methods - practice ) && NOW ( Bi8090 Gene engineering ) && ! Bi5491 Molecular Diagnostics
Basic methods of molecular biology (extraction and purification of DNA, enzymatic reactions, gel electrophoresis) a bioinformatics (manipulating sequence data and pimer design).
Course Enrolment Limitations
The course is only offered to the students of the study fields the course is directly associated with.

The capacity limit for the course is 24 student(s).
Current registration and enrolment status: enrolled: 0/24, only registered: 0/24
fields of study / plans the course is directly associated with
Course objectives
At the end of this course, students should be able to perform methods of DNA cloning, choose suitable vectors and restriction endonucleases for DNA cloning, design primers and use polymerase chain reaction for modification of DNA ends, prepare DNA samples for sequencing, edit and silence genes in vivo using CRISPR/Cas9. Proteins purification.
Learning outcomes
At the end of this course, students should be able to perform methods of DNA cloning, choose suitable vectors and restriction endonucleases for DNA cloning, preparation of molecular probes, design primers and use polymerase chain reaction for modification of DNA ends and prepare DNA samples for sequencing.
Syllabus
  • 1. Cloning of endolysin gene in pUC and pET plasmid vectors (modification of DNA ends by PCR, cloning of PCR products, dephosphorylation of DNA ends by alkaline phosphatase, ligation of vector and foreign DNA by T4 DNA ligase, transformation of competent cells, identification of recombinant clones on ampicillin plates, restriction analysis and construction of restriction maps of recombinant plasmids).
  • 2. Expresion and purification of recombinant protein SH3b_GFP by liquid chromatography.
  • 3. Silencing of septum-coding genes using CRISPR/Cas9 in Escherichia coli. Design of guide RNA for CRISPR editing. Manipulation with dCas9 plasmid. Cassette exchange of insert. Transformation of the cells. Evaluation of the results by light microscopy.
  • 4. Design of plasmid vector using CRISPR-Cas10 system for phage genome editing. The task includes the design of anti-tag sequence, design of a spacer for selection of phage particles. Verification of the functionality of the proposed editing system.
Literature
  • BROWN, T. A. Klonování genů a analýza DNA : úvod. Translated by Martin Fellner. 1. české vyd. V Olomouci: Univerzita Palackého, 2007. xviii, 389. ISBN 9788024417196. info
  • ŠMARDA, Jan, Jiří DOŠKAŘ, Roman PANTŮČEK, Vladislava RŮŽIČKOVÁ and Jana KOPTÍKOVÁ. Metody molekulární biologie (Methods of molecular biology). 1st ed. Brno: Masarykova univerzita, 2005. 194 pp. 1. vydání. ISBN 80-210-3841-1. info
  • SAMBROOK, J., E.F. FRITSCH and T. MANIATIS. Molecular Cloning. A laboratory Manual. Second Edition. Cold Spring Harbor: Cold Spring Harbor Laboratory Press, 1989. ISBN 0-87969-309-6. info
Teaching methods
The course will be held in blocks, in one 3-hour theoretical block, which will be common for both seminar groups and in four 6-hour practical blocks for each seminar group separately (max. 12 students per group). Students will design different plasmid constructs and perform practical experiments based on available protocols.
Assessment methods
The prerequisite for completion of the course is writing the initial theoretical test, submitting the protocols and participation in the practical parts of course.
Language of instruction
Czech
Further comments (probably available only in Czech)
The course is taught annually.
The course is taught: every other week.
Information on course enrolment limitations: Studenti kteří již absolvovali B6492 tento předmět nezapisují z důvodu stejného obsahu. Prosíme striktně o dodržování limitu počtu studentů.
The course is also listed under the following terms Spring 2008 - for the purpose of the accreditation, Spring 2011 - only for the accreditation, Spring 2003, Spring 2004, Spring 2005, Spring 2006, Spring 2007, Spring 2008, Spring 2009, Spring 2010, Spring 2011, Spring 2012, spring 2012 - acreditation, Spring 2013, Spring 2014, Spring 2015, Spring 2016, Spring 2017, spring 2018, Spring 2019, Spring 2020, Spring 2021, Spring 2022.
  • Enrolment Statistics (Spring 2023, recent)
  • Permalink: https://is.muni.cz/course/sci/spring2023/Bi8313