Analiza nadekspresorskih linija uročnjaka (Arabidopsis thaliana L.) za auksin-amidohidrolazu AtILL2

master's thesis

• Title (english): Analysis of auxin amidohydrolase AtILL2 overexpressor lines in Arabidopsis thaliana L.
• Author: Ana Mikelić
• Mentor: Branka Salopek Sondi (mentor)
• Committee member: Aleksandra Maršavelski (predsjednik povjerenstva)
• Committee member: Iva Juranović Cindrić (član povjerenstva)
• Committee member: Davor Kovačević (član povjerenstva)
• Committee member: Morana Dulić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2019-02-27, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry

Abstract

Biljni hormoni auksini imaju važnu ulogu u biljnom rastu i razvitku te u odgovoru biljaka na čimbenike stresa. Aktivan hormon oslobađa se iz konjugata s aminokiselinama djelovanjem enzima auksin-amidohidrolaza koje sudjeluju u homeostazi auksina. Svrha ovog rada bila je istražiti ulogu auksin-amidohidrolaze AtILL2 iz uročnjaka (Arabidopsis thaliana) u odgovoru biljaka na solni i osmotski stres. Analizirane su četiri homozigotne linije A. thaliana ekotipa Ws potencijalnih nadekspresora za fuzijski protein AtILL2 s GFP- i/ili His-privjeskom. Prisutnost transgena provjerena je na razini insercije u genomsku DNA, prisutnosti RNA transkripta i rekombinantnog proteina korištenjem slijedećih metoda: PCR, RT-PCR, SDS-PAGE, afinitetna kromatografija i western-hibridizacijska analiza. U tri linije potvrđena je prisutnost transgena na razini insercije u genomsku DNA i transkripcije u RNA iako nije dokazana prisutnost rekombinantnih proteina niti u jednoj liniji. Testom inhibicije rasta korijena dokazana je povećana otpornost jedne linije na povišeni salinitet i osmolarnost.

Abstract (english)

Phytohormones auxins play an important role in plant growth development and abiotic stress responses. The active auxin may be released from the amino acid conjugates by the action of auxin-amidohydrolase enzymes, which thereby participate in the auxin homeostasis. The aim of this study was to investigate the role of auxin-amidohydrolase AtILL2 from Arabidopsis thaliana in the plant response to salinity and osmotic stress. Four homozygous lines of A. thaliana ecotypes Ws of potential overexpressors for AtILL2 fusion protein with GFP and / or His tags were analyzed. The presence of transgene was verified as insertion into DNA, RNA transcript, and recombinant protein by using PCR, RT-PCR, SDS-PAGE, affinity chromatography and western-hybridization assay. In three lines, the presence of transgene in genomic DNA and RNA transcript was confirmed, although the recombinant protein was not demonstrated in any of the lines. Root growth bioassay confirmed the increased resistance of one line to the examined stressors.

• Keywords: Arabidopsis thaliana; auksin; auksin-amidohidrolaza; linije nadekspresora; osmotski stres; rezistencija; solni stres
• Keywords (english): abiotic stress resistance; Arabidopsis thaliana; auxin; auxin-amidohydrolase; overexpressor line; osmotic stress; salinity stress
• Language: croatian
• URN:NBN: urn:nbn:hr:217:865119
• Study programme: Title: Graduate university studies in chemistry; specializations in: Research programme; Course: Research programme; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra kemije (magistar/magistra kemije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2019-03-13 16:56:29