DNA Manipulation and Synthetic Biology

Code:

B4030

Acronym:

B4030

Keywords
Classification	Keyword
OFICIAL	Biology

Instance: 2013/2014 - 1S

Active?	Yes
Responsible unit:	Department of Biology
Course/CS Responsible:	Master in Cell and Molecular Biology

Cycles of Study/Courses

Acronym	No. of Students	Study Plan	Curricular Years	Credits UCN	Credits ECTS	Contact hours	Total Time
M:BCM	21	Official Study Plan until 2023/2024	1	-	6	42	162

Teaching language

Portuguese

Objectives

Aims of the course unit include: To provide advanced concepts about DNA manipulation, recombinant DNA technologies, cloning and their applications in genetic transformation of organisms and PCR focusing on the molecular techniques involved in the production of recombinant proteins. Provide training in the use of basic bioinformatics tools in analysis and planning of a protein expression project. Primer design and gene optimization. Provide laboratory “hands-on” training in cloning of gene(s) into expression vectors containing different tags and fusion proteins and transformation into appropriate bacterial strains. Familiarization with analysis of original scientific articles, logic of experimental design and data analysis. Training in project work, experimental design and in oral and written communication. Provide the basic concepts and terminology related to synthetic biology.

 

Learning outcomes and competences

Students should understand the rationale for recombinant protein production and understand the underlying molecular processes  of gene expression and regulation. Describe and understand the relevant molecular techniques and DNA manipulation involved in recombinant protein production. Use basic bioinformatics tools in analysis and planning of a protein expression project. Understand gene optimization and perform primer design.

Obtain laboratory skills in gene cloning using various vector types and bacterial cell strains, preparation of plasmid DNA, agarose gel electrophoresis, restriction analysis, primer design and PCR. Analyze original scientific articles, plan experimental design and analyze data. Communicate orally projects and results and write lab reports.

 

Working method

Presencial

Program

Structure and physicochemical properties of DNA. Structure of genes and regulatory sequences. Gene expression and regulation. DNA sequence analysis. Gene optimization. DNA cloning. PCR and PCR based cloning, restriction enzymes. Cloning vectors. Gateway technology. Expression vectors. Vectors containing different tags and fusion proteins. The pET system and the T7 expression system. Regulators of expression (derived from the pLac promoter, expression of the repressor protein in strains with allele lacIq). Bacterial strains with genetic markers for recombinant protein expression. Introduction to Synthetic Biology. Workshop in Synthetic Biology.

Basic bioinformatics tools: database search; BLAST similarity search, restriction analysis, primer design.

Laboratory - Cloning and expression of a gene in E. coli BL21 (DE3) and E. coli Rosetta-Gami 2. Minipreparation of plasmid DNA and its analysis by agarose gel electrophoresis. Isolation of genes by PCR with primers with RE adapters. Restriction of plasmids and PCR products. Insertion of the gene under study in the appropriate vector(ex. pET30a). Transformation into competent E. coli BL21 (DE3) or E. coli Rosetta-Gami 2, and plating on media containing appropriate antibiotics and IPTG. Screening of recombinants by PCR. Preliminary study of bacterial extracts by western blot.

 

 

Mandatory literature

NCBI - National Center for Biotechnology Information; A Science Primer - http://www.ncbi.nlm.nih.gov/About/primer/, NCBI

Complementary Bibliography

Videira Arnaldo 340; Engenharia genética. ISBN: 9789727577439

Teaching methods and learning activities

Students are encouraged to engage in projects, both in working groups or individually, and to take the initiative to investigate specific topics. The use of real examples of research and "case studies", provide the context of reality and allows the acquisition of the notion that science is a continuous search for answers to questions that are not ever completely resolved. Activities include lectures, discussion groups, tutorials, problem solving, debates, etc. Emphasis also will be placed in wet lab training.

 

Assessment tools include presentation and discussion of original scientific articles, a written laboratory report on the project developed and a final exam.

keywords

Natural sciences > Biological sciences > Biology > Molecular biology
Natural sciences > Biological sciences > Biological engineering > Genetic engineering

Evaluation Type

Distributed evaluation with final exam

Assessment Components

designation	Weight (%)
Exame	50,00
Participação presencial	0,00
Teste	15,00
Trabalho escrito	35,00
Total:	100,00

Eligibility for exams

Compulsory class attendance (minimum 2/3) and submission of Laboratory class report.

Calculation formula of final grade

Final Exam (10 points) + Report laboratory work (7 points) + Oral presentation of laboratory work (3 points).

Classification improvement

Final Exam (10 points)