Molecular biology (2017/2018)



Course code
4S00800
Credits
12
Coordinator
Massimiliano Perduca
Academic sector
BIO/11 - MOLECULAR BIOLOGY
Language of instruction
Italian
Teaching is organised as follows:
Activity Credits Period Academic staff Timetable
teoria 9 II sem. Massimiliano Perduca

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laboratorio [1° turno] 3 II sem. Massimiliano Perduca

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laboratorio [2° turno] 3 II sem. Massimiliano Perduca

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Learning outcomes

The aim of this course is to give the students the basic knowledge of the molecular mechanisms concerning transmission, variation and expression of the genetic information.
At the end of the course the students will be able to recognize the major molecular mechanisms of life in prokaryotic and eukaryotic cells.

Syllabus

Theory:
> Genetic information and informational molecules
General introduction and historical hints. The chemical structure of DNA and RNA. Three dimensional structure of DNA. Physico-chemical properties of DNA.
> Molecular Biology techniques
Agarose gel electrophoresis. Nucleic acid hybridization. Polymerase chain reaction (PCR). Restriction endonucleases. Cloning and sub-cloning. gene expression systems.
> DNA, RNA and gene structure
Definition of gene coding and regulatory regions. From genes to proteins; messenger RNA, transfer RNA and ribosomal RNA.
> Genome organization and evolution
DNA content and number of genes. Mutations, DNA rearrangement and genome evolution. The organelle genomes. Interrupted genes; introns. cDNA. Gene families and duplication. DNA repeats.
> Transposable elements
Transposition mechanisms and control. Retroviruses and retrotransposones. Transposons.
> Chromatin and chromosomes
Nucleosomes, histones and their modifications. Higher organization levels of chromatin. Heterochromatin and euchromatin. Eukaryotic chromosomes, telomeres and centromeres.
> DNA replication
DNA polymerases. Proofreading activity of DNA polymerases. Replication mechanism in bacteria and eukaryotic cells.
> Introns and RNA splicing
Features of spliceosomal introns. Spliceosome and splicing mechanism. Alternative splicing and trans-splicing. Other kinds of introns: group I and group II introns and tRNA introns. The intron movement. RNA editing. Ribozymes and riboswitch.
> DNA mutation and repair
Spontaneous mutations and mutations caused by physical and chemical mutagens. Pre- and post-replicative repair systems. Recombination in the immunity system cells. Approaches to homologous recombination.
> Regulation of gene expression
Bacterial promoters. The operon. Activators, repressors and coactivators. Signal transductions and two component regulation systems. Eukaryotic promoters. Activators, repressors and coactivators. Gene expression and chromatin modifications. Epigenetic mechanisms.
> RNAs and transcription
Different types of RNA: synthesis and maturation. Bacterial RNA polymerase. Sigma factors. Eukaryotic RNA polymerases. Eukaryotic mRNAs: capping, polyadenylation, cytoplasmic localization. The transcription process in bacteria and in eukaryotic cells.
> Translation
Ribosomes. tRNA structure and function. Aminoacyl-tRNA synthesis. Initiation in bacteria and eukaryotic cells. Polypeptide chain synthesis and translation end. Regulation of translation.
> Protein localization.

One credit of the course (corresponding to 8 hours) will be kept for the students to discuss an important topic chosen from the research literature in Molecular Biology.

Introduction to the Laboratory Course:
> Nucleic acids isolation: basis, comparison of several extraction protocols, nucleic acids isolation troubleshooting.
> Nucleic acids electrophoresis: agarose gels, polyacrylamide gels, denaturing and non-denaturing gels.
> Spectrophotometric quantitation of isolated nucleic acids.
> PCR
1.What is PCR?
2. Reagents: efficiency, specificity, fidelity
3. PCR cycle. Final number of copies of the target sequence
4.Amplifying the correct product: detection and analysis of PCR products, how to avoid contamination.
5. Techniques and applications.

Practical:
Subcloning and protein expression:
PCR
Electrophoresis and DNA gel extraction
Plasmidic DNA purification
Digestion with restriction enzymes and ligation
Transformation and colony PCR
Heterologous expression of a recombinant protein in E. coli and analysis by SDS PAGE and Western Blot.

Assessment methods and criteria

Oral examination.
The final exam will be on three questions concerning any of the topics treated during the course and it is passed if both answers are positive.

Reference books
Activity Author Title Publisher Year ISBN Note
teoria Jocelyn E. Krebs, Elliott S. Goldstein, Stephen T. Kilpatrick Lewin's Genes XII (Edizione 12) Jones & Bartlett Pub 2017 1284104494
teoria Bruce Alberts, Alexander Johnson, Julian Lewis, David Morgan, Martin Raff Molecular Biology of the Cell (Edizione 6) Garland Science 2014 0815344643
teoria Nancy Craig, Rachel Green, Carol Greider, Gisela Storz, Cynthia Wolberger, Orna Cohen-Fix Molecular Biology: Principles of Genome Function (Edizione 2) OUP Oxford 2014 0199658579
teoria Arnold Berk, Chris A. Kaiser, Harvey Lodish, Angelika Amon, Hidde Ploegh, Anthony Bretscher, Monty Krieger, Kelsey C. Martin Molecular Cell Biology (Edizione 8) WH Freeman 2016 1464187444
teoria Geoffrey M. Cooper, Robert E. Hausman The cell: a molecular approach (Edizione 6) Sinauer Associates, Inc 2013 978-1-60535-155-1
laboratorio Erik Pierre Molecular Cloning (Edizione 1) Ml Books International 2015 1632394685
laboratorio Michael R. Green e Joseph Sambrook Molecular Cloning: A Laboratory Manual, Fourth Edition (Edizione 4) CSHL Press 2012 978-1-936113-42-2
laboratorio Erik Pierre Molecular Cloning (Edizione 1) Ml Books International 2015 1632394685
laboratorio Michael R. Green e Joseph Sambrook Molecular Cloning: A Laboratory Manual, Fourth Edition (Edizione 4) CSHL Press 2012 978-1-936113-42-2
Teaching aids
Title Format (Language, Size, Publication date)
10-The ribosomal A-site finger  pdfpdf (en, 2412 KB, 10/04/18)
11-Structure and function of the N-terminal domain of the yeast telomerase reverse transcriptase  pdfpdf (en, 3515 KB, 10/04/18)
12-End resection by RecBCD during double-stranded DNA break repair  pdfpdf (en, 4416 KB, 10/04/18)
13-APE2 promotes DNA damage response pathway  pdfpdf (en, 4389 KB, 10/04/18)
14-RecA requires two molecules of Mg2+ ions  pdfpdf (en, 2355 KB, 10/04/18)
15-Genome-wide dose-dependent inhibition of histone deacetylases  pdfpdf (en, 10234 KB, 10/04/18)
16-Mechanisms of improved specificity of engineered Cas9s  pdfpdf (en, 3291 KB, 10/04/18)
17-Recognition of DNA binding site by Integration Host Factor  pdfpdf (en, 2905 KB, 10/04/18)
1-BCL11A Controls the Fetal to Adult Hemoglobin Switch  pdfpdf (en, 5588 KB, 10/04/18)
2-Ribothrypsis, a novel process of canonical mRNA decay  pdfpdf (en, 1716 KB, 10/04/18)
3-I-Motif of cytosine-rich human telomere DNA  pdfpdf (en, 4476 KB, 10/04/18)
4-Structure of a eukaryotic cytoplasmic pre‐40S ribosomal subunit  pdfpdf (en, 2304 KB, 10/04/18)
5-Structural and functional analysis of ribosome assembly factor Efg1  pdfpdf (en, 8093 KB, 10/04/18)
6-Spliceosomal protein U1A is involved in alternative splicing  pdfpdf (en, 6006 KB, 10/04/18)
7-Recruitment of UvrBC complexes to UV-induced damage  pdfpdf (en, 1721 KB, 10/04/18)
8-RNA polymerase II-associated TFIIF-like complex  pdfpdf (en, 5027 KB, 10/04/18)
9-Global delay in nascent strand DNA methylation  pdfpdf (en, 2214 KB, 10/04/18)
Gruppi Esposizione Articolo Biotecnologie 17-18  pdfpdf (it, 14 KB, 08/03/18)
Laboratorio Didattico Biologia Molecolare I Turno 17-18  pdfpdf (it, 10 KB, 08/03/18)
Laboratorio Didattico Biologia Molecolare II Turno 17-18  pdfpdf (it, 9 KB, 08/03/18)

STUDENT MODULE EVALUATION - 2017/2018