Usporedba metoda za obilježavanje i detekciju pri hibridizaciji DNA po Southern-u

undergraduate thesis

• Title (english): Comparison of methods for DNA labelling and detection used in Southern blot hybridization
• Author: Maša Lač
• Mentor: Anamarija Štafa (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: 2020-07-10, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Hibridizacija po Southern-u koristi se za detekciju sekvencija DNA, a temelji se na komplementarnom sparivanju (hibridizaciji) jednolančane obilježene DNA sonde s jednolančanom ciljnom sekvencijom DNA na membrani. Hibridizacija se odvija pri uvjetima koji omogućuju komplementarno sparivanje DNA sonde s ciljnom regijom, uz minimalnu količinu nespecifičnog vezanja sonde na membranu. DNA sonde mogu se obilježavati radioaktivno, koristeći radioizotope (³²P, ³⁵S, ¹²⁵I, ³H) ili neradioaktivno, koristeći specifične molekule poput biotina ili digoksigenina (DIG). Najčešće korištena metoda za detekciju radioaktivno obilježenih sondi je autoradiografija, dok se detekcija neradioaktivno obilježenih sondi temelji na kemiluminiscencijskim i kromogenim reakcijama. U ovom radu uspoređene su različite metode obilježavanja i detekcije pri hibridizaciji po Southern-u te su navedene njihove prednosti i mane.

Abstract (english)

Southern blot is a method used for detection of specific DNA sequences and it is based on the complementary pairing of a labelled single-stranded DNA probe with target single-stranded DNA bound to the membrane. Hybridization occurs under stringent conditions which allow specific binding of a probe with a target sequence with the minimal nonspecific binding of DNA probe to the membrane. Probes can be labelled radioactively, using radioisotopes (³²P, ³⁵S, ¹²⁵I, ³H) or non-radioactively, using specific biotin or digoxigenin (DIG) molecules. Most commonly used detection method for radiolabeled probes is autoradiography, while detection of non-radioactive methods is based on chemiluminescence and chromogenic reactions. In this thesis, different methods for labelling and detection during Southern blot hybridization were compared and their advantages and disadvantages are indicated.

• Keywords: detekcija DNA; hibridizacija po Southern-u; obilježavanje DNA
• Keywords (english): detection of DNA; labelling DNA; Southern blot
• Language: croatian
• URN:NBN: urn:nbn:hr:159:161534
• 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: 2020-07-27 09:35:02