Generation of Arabidopsis thaliana (Heynh.) plants containing the TROL protein labeled with HA and FLAG protein markers

master's thesis

• Title: Transgenične biljke Arabidopsis thaliana (Heynh.) koje eksprimiraju protein TROL obilježen proteinskim biljezima HA i FLAG
• Author: Jelena Kovačević
• Mentor: Hrvoje Fulgosi (mentor)
• Committee member: Nataša Bauer (predsjednik povjerenstva)
• Committee member: Hrvoje Fulgosi (član povjerenstva)
• Committee member: Željka Vidaković-Cifrek (član povjerenstva)
• Committee member: Dijana Škorić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2013-02-05, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Transformacija biljaka metodom “floral dip” postupak je unosa T-DNA sekvence u genom stanica ovarija prije oplodnje. U ovom radu opisala sam transformaciju biljaka Arabidopsis thaliana molekulom T-DNA iz plazmidnog vektora pH7WG2,0 TROL-HA-FLAG. TROL je transmembranski protein koji sudjeluje u regulaciji puta prijenosa elektrona na tilakoidnim membranama kloroplasta. Dodatak proteinskih biljega HA i FLAG na protein TROL olakšava njegovu imunodetekcijsku provjeru i omogućuje daljnja istraživanja funkcije proteina TROL. Kako bih dokazala uspješnost transformacije biljaka genom za protein TROL-HA-FLAG, biljke sam analizirala metodama selekcije sjemenki na hranjivoj podlozi s antibiotikom, PCR provjere genoma i imunodetekcije proteina u listovima odraslih biljaka. U transformiranim biljkama sam imunodetekcijom potvrdila lokalizaciju proteina TROL-HA-FLAG u tilakoidnim membranama kloroplasta. Daljnja istraživanja omogućit će detaljniju potvrdu funkcionalnosti proteina TROL-HA-FLAG.

Abstract (english)

Floral dip is a transformation method that results in the insertion of T-DNA into the genome of unfertilized ovary cells. This thesis describes the transformation of Arabidopsis thaliana plants with T-DNA from pH7WG2,0 TROL-HA-FLAG plasmid vector. TROL is a transmembrane protein that regulates electron transport flow on the thylakoid membranes of chloroplasts. Addition of protein tags HA and FLAG to the TROL protein allows easier immunodetection analysis and enables further studies of the plant TROL protein function. In order to show the success of plant transformation with the TROL-HA-FLAG protein gene, the plants were subjected to multiple analyses such as seed selection method on growth plates with an added antibiotic, plant DNA PCR analysis and immunodetection analysis of plant leaf proteins. Due to the immunodetection analysis, I managed to confirm the localization of TROL-HA-FLAG protein in chloroplast thylakoid membranes of transformed plants. Further research could better show if TROL-HA-FLAG protein function is diminished.

• Keywords: „floral dip“; transformacija; HA epitop; fotosinteza; selekcija
• Keywords (english): floral dip; transformation; HA tag; photosynthesis; selection
• Language: croatian
• URN:NBN: urn:nbn:hr:217:496612
• Study programme: Title: Molekularna biologija; Study programme type: sveučilišni; Study level: diplomski; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Restricted access
• Created on: 2017-04-27 09:26:49