Optimizacija modularnog sustava ekspresijskih vektora za primjenu CRISPR/Cas9 tehnologije u kulturi animalnih stanica

master's thesis

• Title (english): Optimization of a modular system of expression vectors for the application of CRISPR/Cas 9 technology in animal cell culture
• Author: Bruno Bračun
• Mentor: Vlatka Čubrić Čurik (mentor)
• Committee member: Vlatka Čubrić Čurik (predsjednik povjerenstva)
• Committee member: Ino Čurik (član povjerenstva)
• Committee member: Maja Ferenčaković (član povjerenstva)
• Granter: University of Zagreb; Faculty of Agriculture; (Animal Science); Zagreb
• Defense date and country: 2021-09-20, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Agronomy

Abstract

CRISPR/Cas9 modularni sustav je sustav koji se koristi za epigenetičku manipulaciju, odnosno za aktivaciju ili deaktivaciju genske transkripcije putem metilacije, odnosno demetilacije CpG otoka u individualnim promotorima i acetilacije i deacetilacije krajeva histonskih repova. No, za samu ekspresiju modularnog sustava, najprije je potrebno konstruirati plazmid, koji se zatim prenosi u eukariotsku stanicu. Takav vektor trebao bi biti najmanji mogući tj. trebao bi sadržavati samo sekvence koje se sastoje od komponenate koje čine CRISPR/Cas9 modularni sustav. Takva zadaća omogućena je konstrukcijom minimalnog plazmida koji će biti optimiziran koristeći najbolje krajeve za sklapanje metodom Golden Gate, svih potrebnih dijelova koje će plazmid u eukariotskoj stranici eksprimirati. Dobiveni podaci su potvrdili uspješnost sklapanja minimalnog plazmida sa krajevima koji će omogućiti daljnju uspješnu manipulaciju konstruktom korištenim u CRISPR/Cas9 tehnologiji.

Abstract (english)

The CRISPR/Cas9 modular system is a system for epigenetic manipulation, i.e., activation and deactivation of gene transcription by metilation or demetilation of CpG islands in individual promoter regions or by acetilation and deacetilation of the remaining histone tails. For expression of the modular system itself, a plasmid must first be made to be transfected into the eukaryotic cell. This system should be as small as possible, with sequences containing only parts that make up the modular CRISPR/Cas9 system for epigenetic manipulation. Such a mission would only be successful by making the smallest possible plasmid, optimized by the best possible overhangs for assembling all the parts that this plasmid will express in the eukaryotic cell. Recieved data have shown success in folding minimal plasmid with ends that will allow further succes with manipulation of construct used in CRISPR/Cas9 technology.

• Keywords: CRISPR/Cas9; plazmid; epigenetička regulacija
• Keywords (english): CRISPR/Cas9; plasmid; epigenetic regulation
• Language: croatian
• URN:NBN: urn:nbn:hr:204:784953
• Study programme: Title: Animal Genetics and Breeding; Study programme type: university; Study level: graduate; Academic / professional title: magistar/ magistra inženjer/inženjerka genetike i oplemenjivanja životinja (magistar/ magistra inženjer/inženjerka genetike i oplemenjivanja životinja)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2021-11-24 09:59:26