Plant molecular technologies (2021/2022)
Scientific Disciplinary Sector (SSD)
BIO/04 - PLANT PHYSIOLOGY
The teaching is organized as follows:
At the end of the course, the student will acquire the theoretical and practical basis for the design of various types of genetic constructs aimed at developing transgenic, cisgenic, and intragenic plants. In addition, genome editing tools will be covered. Attention will be paid to the application of these strategies for qualitative and quantitative crop improvement.
Molecular mechanism of stable transformation via Agrobacterium: current knowledge of the process of T-DNA integration into the plant genome; transgene integration, stability, methylation and silencing; strategies to avoid transgene silencing; promoters used for genetic construct preparation (constitutive, spatiotemporal, inducible, synthetic); cis–trans engineering; synthetic circuits (e.g. hormometer); multigene engineering; artificial miRNA and target mimicry; cisgenesis and intragenesis; cisgenic and intragenic genetic constructs; strategies to remove marker genes from transgenic plants; artificial programmable DNA nucleases (ZFNs and TALENs) and RNA-guided DNA nucleases for genome engineering; type II CRISPR-Cas9 mechanism of action; prime editing; single-base editing; sgRNA design; minimization of off-target activity; dead Cas9 and nickase Cas9: application of the CRISPR-Cas system beyond genome editing; (CRISPR)-based tissue-specific knockout system; genetic constructs for genome engineering using the CRISPR-Cas9 system, screening of mutants generated by CRISPR system.
Supplied educational material:
Power point lessons, relevant research articles and reviews.
1) Design of guide RNAs employing bioinformatics software, in vitro transcription of guide RNAs, identification of sgRNAs most effective in inducing Cas9-mediated cutting of target DNA fragments in vitro; molecular analysis of transgenic tomato plants previously transformed with the gene construct of interest for editing in the target gene, amplification and sequencing of the target region, analysis of results.
The purpose of the exam is to ascertain the student's knowledge of the topics covered during the lessons and workshops. The exam will consist of an individual written test, containing 3 open-ended questions. In addition, each student will present at the end of the lectures a scientific article, chosen from a list of articles proposed by the teacher. The evaluation criteria for the presentation of the article will concern the general comprehension, the deepening of the topic at the state of the art level, the property of language, the ability to analyze and critically discuss the results. The final grade, expressed in thirtieths, will be given considering one third for the presentation of the article and two thirds for the written test. The method of examination will be the same for attending and non-attending students.