Optimizacija metod kolokalizacije za funkcionalno analizo genov krompirja, ki sodelujejo pri obrambnem odzivu na krompirjev virus Y

Race, Andrej (author), Gruden, Kristina (mentor)

PVY predstavlja najbolj ekonomsko škodljiv virus za gojenje krompirja. Precej podatkov o mehanizmih interakcije med PVY in krompirjem je že poznanih, od interakcij različnih sevov PVY z različnim genotipi krompirja, do vpliva hormonov na imunski odgovor krompirja in vlog posameznih genov PVY ter krompirja v tej interakciji. Namen tega dela je bilo preverjanje interakcije med proteini imunskega odgovora krompirja s pomočjo metode kolokalizacije za interakcije, ki so bile potrjene kot pozitivne z metodo dvohibridnega sistema kvasovk. V eksperimentu smo najprej pripravili dva seva transgenih agrobakterij, od katerih je en vseboval binarni plazmid z zapisom za ERF98.2-GFP in drugi binarni plazmid z zapisom za PP2C-GFP. Agrobakterije smo nato uporabili za analizo lokalizacije SAPK8-RFP, ERF98.2-GFP in PP2C-GFP v listih N. benthamiana in preverili, ali pride do kolokalizacije SAPK8-RFP in ERF98.2-GFP ter SAPK8-RFP in PP2C-GFP. Rezultati so pokazali, da je do kolokalizacije SAPK8-RFP in ERF98.2-GFP prišlo v jedru in citoplazmi, medtem ko je do kolokalizacije SAPK8-RFP in ERF98.2-GFP prišlo le v jedru, čeprav v večini celic ni bila vidna.

• Language: Slovenian
• Keywords: krompir-PVY interakcija, kolokalizacija, SAPK8, ERF98.2, PP2C
• Work type: Bachelor thesis/paper
• Typology: 2.11 - Undergraduate Thesis
• Organization: FKKT - Faculty of Chemistry and Chemical Technology
• Year: 2019
• PID: 20.500.12556/RUL-110324
• COBISS.SI-ID: 1538426563
• Publication date in RUL: 13.09.2019

Secondary language

• Language: English

Abstract

PVY is the most economically harmful virus for potato growing. The relationship between PVY and potatoes was so far intensively studied, describing the interactions of different PVY strains with different potato cultivars, the influence of hormones on the potato immune response, the roles of individual PVY and the potato genes in that interaction. The purpose of this work was to verify the interaction between the proteins of the potato immune response using the method of co-localisation for interactions that were positive in the yeast two-hybrid system. In the experiment, transgenic agrobacteria were first prepared using two plasmids, one containing a binary plasmid with an insert coding for ERF98.2-GFP and the other containing a binary plasmid with an insert coding for PP2C-GFP. Agrobacteria were then used to analyze the localization of SAPK8-RFP, ERF98.2-GFP, and PP2C-GFP in N. benthamiana leaves and to check whether co-localization occurred between SAPK8-RFP and ERF98.2-GFP, as well as SAPK8-RFP and PP2C-GFP. The results showed that the first co-localization occurred in the nucleus and cytoplasm, while the second co-localization occurred in the nucleus, although it was not visible in most cells.

• Keywords: potato-PVY interaction, co-localization, SAPK8, ERF98.2, PP2C