Usporedba različitih postupaka izdvajanja virusne RNA iz tkiva ptičjeg mozga

master's thesis

• Title (english): Comparison of different methods for extraction of viral RNA from avian brain tissue
• Author: Iva Podgorski
• Mentor: Vladimir Savić (mentor)
• Mentor: Inga Marijanović (mentor)
• Committee member: Inga Marijanović (predsjednik povjerenstva)
• Committee member: Vladimir Savić (član povjerenstva)
• Committee member: Martina Šeruga Musić (član povjerenstva)
• Committee member: Dijana Škorić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2012-07-03, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Brza i točna dijagnostika zaraznih bolesti životinja vrlo je bitna radi pravodobnog sprječavanja širenja bolesti i posljedičnih velikih gubitaka. Tehnike molekularne biologije, posebno izdvajanje RNA i DNA, nalaze veliku primjenu u dijagnostici virusnih infekcija. U ovom radu je istražen učinak uranjanja uzoraka u tekući dušik i uspješnost 4 različite metode izdvajanja virusne RNA iz tkiva mozga pokusno zaraženih pilića koje je pohranjeno na -70°C. RNA je izdvojena korištenjem komercijalnih reagensa: 1) Roche High Pure Viral Nucleic Acid Kit, 2) TRIzol®, 3) Qiagen RNeasy Lipid Tissue Mini Kit te 4) kombinacijom TRIzol® i Roche High Pure Viral Nucleic Acid Kita. Prinosi izdvojene RNA su uspoređeni RRT-PCR analizom na uređaju LightCycler 1.5. Svaka metoda je dala dovoljnu količinu RNA za potvrdu prisutnosti ptičjeg paramiksovirusa tipa 1, ali najbolji prinosi dobiveni su korištenjem komercijalnog kompleta Qiagen RNeasy Lipid Tissue Mini Kit i korištenjem reagensa TRIzol®. Uranjanje tkiva u tekući dušik nije imalo utjecaja na prinos RNA.

Abstract (english)

Rapid and accurate diagnosis of animal infectious diseases is cruical for timely prevention of the spread and the consequent heavy losses. Molecular biology techniques, especially RNA and DNA isolation, are extensively used in viral infection diagnosis. This study investigates the effect of sample freezing in liquid nitrogen and viral RNA yield by use of four different RNA isolation methods from experimentally infected chickensʼ brain tissue previously stored at -70°C. RNA was isolated by using commercial reagents: 1) Roche High Pure Viral Nucleic Acid Kit, 2) TRIzol®, 3) Qiagen RNeasy Lipid Tissue Mini Kit and 4) combination of TRIzol® and Roche High Pure Viral Nucleic Acid Kit. Yields of isolated RNA were compared by RRT-PCR in LightCycler 1.5. Each method provided enough RNA to detect avian paramyxovirus type 1, but isolation by using Qiagen RNeasy Lipid Tissue Mini Kit and TRIzol® gave the best RNA yield. Freezing of the brain tissue in liquid nitrogen did not influence the yield of RNA.

• Keywords: ptičji paramiksovirus tipa 1; newcastelska bolest; RRT-PCR
• Keywords (english): avian paramyxovirus type 1; Newcastle disease; RRT-PCR
• Language: croatian
• URN:NBN: urn:nbn:hr:217:950519
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Access restricted to students and staff of home institution
• Created on: 2017-05-02 10:29:59