Optimizacija lančane reakcije polimerazom za umnažanje mikrosatelitnih biljega divlje mačke Felis silvestris Schreber, 1777

undergraduate thesis

• Title (english): Optimization of polymerase chain reaction for amplification of microsatellite markers of wildcat Felis silvestris Schreber, 1777
• Author: Lucija Jaša-Šangulin
• Mentor: Ana Bielen (mentor)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Biochemical Engineering); (Laboratory for Biology and Microbial Genetics); Zagreb
• Defense date and country: 2017-07-17, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Genomska ĉistoća populacije divlje maĉke u mnogim je europskim zemljama ugrožena zbog uĉestale hibridizacije s domaćom maĉkom. S ciljem utvrĊivanja stanja populacije u Hrvatskoj potrebno je provesti genotipizaciju divljih i domaćih maĉaka pomoću umnažanjem mikrosatelitnih biljega lanĉanom reakcijom polimerazom. U ovom je radu provedena optimizacija lanĉane reakcije polimerazom za umnažanje 12 odabranih mikrosatelitnih biljega divlje maĉke. UtvrĊena je optimalna koliĉina DNA kalupa od 10 ng i koncentracija oligonukleotidnih poĉetnica od 0.2 μM. Optimizirani su i uvjeti provoĊenja reakcije; temperatura vezanja poĉetnica od 57°C i vremena vezanja poĉetnica i produljenja lanaca od 90 sekundi. Konaĉno, opisani optimalni uvjeti prilagoĊeni su provedbi multipleks PCR-a radi uštede vremena i resursa. Rezultati ovog rada će u nastavku istraživanja poslužiti za umnažanje mikrosatelitnih biljega većeg broja divljih i domaćih maĉaka s ciljem odreĊivanja stupnja hibridizacije s domaćim maĉkama u Republici Hrvatskoj.

Abstract (english)

Genomic purity of European wildcat population is compromised by frequent hybridisation with domestic cats. Since the state Croatian wildcat population is unknow, it is necessary to carry out genotyping, i.e. amplify the microsatellite markers by polymerase chain reaction. In this work, optimization of PCR for amplification of 12 selected wildcat microsatellite markers was performed. An optimal amount of 10 ng DNA template and 0.2 μM of oligonucleotide primers was determined.Thermal cycling conditions were also optimized; optimal annealing temperature was 57°C and annealing and elongation time of 90 seconds. Finally, reaction conditions were further adjusted to perform multiplex PCR in order to save time and resources. Results of this work will be the basis of future amplification of microsatellite markers of wild and domestic cats in order to determine genomic purity of wildcat population in Croatia.

• Keywords: Felis silvestris Schreber 1777; mikrosatelitni biljezi; optimizacija PCR reakcije
• Keywords (english): Felis silvestris Schreber 1777; microsatellite markers; optimization; PCR
• Language: croatian
• URN:NBN: urn:nbn:hr:159:816388
• Study programme: Title: Biotechnology; Study programme type: university; Study level: undergraduate; Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-07-24 11:35:54