Molekularna karakterizacija autohtonih sojeva Sinorhizobium meliloti izoliranih iz različitih regija Hrvatske

master's thesis

• Author: Edita Hajdari
• Mentor: Sanja Sikora (mentor)
• Committee member: Sanja Sikora (predsjednik povjerenstva)
• Committee member: Snježana Bolarić (član povjerenstva)
• Committee member: Dubravko Maćešić (član povjerenstva)
• Committee member: Sanja Kajić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Agriculture; (Department of Microbiology); Zagreb
• Defense date and country: 2016-09-28, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Agronomy

Abstract

Simbiozni odnos između biljke domaćina – lucerne (Medicago sativa L.) i kvržične bakterije vrste Sinorhizobium meliloti smatra se vrlo značajnim u održivoj poljoprivrednoj proizvodnji. Simbiozna fiksacija dušika osigurava unos značajnih količina dušika u poljoprivredno tlo, što omogućuje smanjenu primjenu mineralnih dušičnih gnojiva. Ostvarivanje ovakvih odnosa moguće je osigurati predsjetvenom bakterizacijom sjemena leguminoza, gdje se kao inokulum koriste visoko učinkoviti sojevi kvržičnih bakterija poznatih karakteristika. Osim prisustva nekog drugog izvora dušika, uspješnost bakterizacije može reducirati i prisutnost autohtonih kompetitivnih sojeva rizobija u tlu. Takvi sojevi bakterija mogu biti prilagođeni uvjetima u tlu, inficirati prisutnu biljku domaćina, ali nije nužno da dolazi do fiksacije dušika ili da su fiksirane dovoljne količine. Zbog navedenog je važna izolacija i karakterizacija autohtonih sojeva S.meliloti, što je i bio glavni cilj ovoga istraživanja. Autohtoni sojevi izolirani su iz tla prikupljenog s područja Istarske, Zadarske, Koprivničko – križevačke i Brodsko – posavske županije. Istraživanjem je obuhvaćeno 20 autohtonih sojeva, referentni soj S. meliloti 2011 te tipski soj Sinorhizobium medicae 18864. Identifikacija izolata na razini vrste provedena je primjenom 16S rDNA PCR – RFLP metode, nakon koje su reducirani izolati za koje nije određena pripadnost vrsti S. meliloti. Utvrđeno je da 14 od 20 autohtonih izolata pripada navedenoj vrsti, dok se ostalih 6 značajno razlikovalo. Za identifikaciju izolata na razini soja korištene su RAPD – PCR i ERIC – PCR metode. Identifikacijom ovih izolata pomoću navedenih metoda utvrđena je značajna genetska varijabilnost unutar prirodnih populacija S. meliloti. Potrebna je daljnja fenotipska i simbiozna karakterizacija ovih sojeva kako bi se mogli selekcionirati najkvalitetniji sojevi za potencijalnu primjenu u uzgoju lucerne.

Abstract (english)

The symbiotic relationship between host plant – alfalfa (Medicago sativa L.) and nodule bacteria Sinorhizobium meliloti is considered to be very significant in sustainable agricultural production. Symbiotic nitrogen fixation provides input of significant nitrogen amounts in agricultural soil, which enable reduced application of mineral nitrogen fertilizers. The realization of these relations is possible by legumes seed inoculation, where are highly effective strains of rhizobia, with well known characteristics, used as an inoculum. Except of presence of some other nitrogen sources, the bacterization efficacy can be reduced also by presence of competitive indigenous rhizobial strains in soil. Such strains can be adapted to the all conditions in soil, infect the present host plant, but it is not necessarily that the process of nitrogen fixation will happen, or that sufficient quantity of nitrogen will be fixed. That is why the isolation and characterization of indigenous strains of S. meliloti is so important, and that was also the objective of this study. Indigenous strains were isolated from the soil collected in the Istrian, Zadar, Koprivnica – Križevci and Brod – Posavina county. The study includes 20 indigenous strains, one reference strain S. meliloti 2011 and one standard strain Sinorhizobium medicae 18864. Identification to the species level was performed by 16S rDNA PCR – RFLP method, and after that isolates which were not S. meliloti species were reduced. It has been found that 14 of 20 indigenous strains belongs to the specified species, while the other 6 were significantly different. Identification to the strain level was performed by RAPD – PCR and ERIC – PCR methods. Through identifications of these isolates done by specified methods, significant genetic variability was established within natural populations of S. meliloti. Further fenotipic and simbiotic characterizations of this strains are required in order to select the strains with highest qualities for potential use in alfalfa cultivation.

• Keywords: simbiozna fiksacija dušika; lucerna (Medicago sativa L.); autohtoni sojevi Sinorhizobium meliloti; 16S rDNA PCR – RLFP; RAPD – PCR; ERIC – PCR
• Keywords (english): symbiotic nitrogen fixation; alfalfa (Medicago sativa L.); indigenous strains of Sinorhizobium meliloti; 16S rDNA PCR – RFLP; RAPD – PCR; ERIC – PCR
• Language: croatian
• URN:NBN: urn:nbn:hr:204:414731
• Study programme: Title: Agroecology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/ magistra inženjer/inženjerka agroekologija (magistar/ magistra inženjer/inženjerka agroekologija)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Access restricted to students and staff of home institution
• Created on: 2016-11-29 08:07:04