Vzpostavitev modelnega sistema za proizvodnjo nanoteles v kvasovki Saccharomyces cerevisiae

diplomsko delo

Žan Žnidar (Author), Uroš Petrovič (Mentor)

Abstract

Enodomenska protitelesa (nanotelesa) so v zadnjem desetletju vse bolj priljubljena, zahvaljujoč stabilnosti in enostavni strukturi. Ker sestojé iz zgolj ene polipeptidne verige, jih je možno pravilno zvite pripraviti v bakterijah ali kvasovkah, zato lahko ustvarimo produkcijske sisteme za proizvodnjo, ki so relativno poceni in dostopni. Kvasovka Saccharomyces cerevisiae sodi v skupino varnih organizmov (GRAS), zato so njihovi proteinski produkti široko uporabni – tudi v farmacevtiki. Kvasovke smo transformirali s plazmidom, ki je v vključku vseboval z zapisom za signalni peptid a faktorja spojen zapis za nanotelo proti proteinu bodice virusa SARS CoV 2, inducirali izražanje pod nadzorom inducibilnega promotorja, nato pa iz gojišča izolirali nanotelo. Protein se je uspešno izražal in izločal v gojišče, kar smo ovrednotili s prenosom western. V tej diplomski nalogi smo dokazali, da je predlagani modelni sistem primeren za proizvodnjo izbranega nanotelesa.

Keywords

kvasovke;Saccharomyces cerevisiae;proizvodnja proteinov;nanotelesa;sekrecija;diplomska dela;

Data

Language:	Slovenian
Year of publishing:	2023
Typology:	2.11 - Undergraduate Thesis
Organization:	UL FKKT - Faculty of Chemistry and Chemical Technology
Publisher:	[Ž. Žnidar]
UDC:	577.112:577.27(043.2)
COBISS:	165400835
Views:	51
Downloads:	11
Average score:	0 (0 votes)
Metadata:

Other data

Secondary language:	English
Secondary title:	Creating a model system for production of nanobodies in yeast Saccharomyces cerevisiae
Secondary abstract:	Single-domain antibodies (nanobodies) have gained popularity in the last decade due to their stability and simple structure. Consisting of only one polypeptide chain, they can be synthesized by bacteria or yeasts as recombinant proteins, and still fold correctly. This allows us to create production systems which are relatively cheap and accessible. Yeast Saccharomyces cerevisiae belongs to GRAS organisms, thus allowing their protein products to have a broad range of applications – including in pharmacy. Upon transforming yeast cells with a plasmid containing a gene for a nanobody against the SARS CoV 2 spike protein following a gene for the alpha-factor leader peptide, the production under the control of an inducible promoter was induced, then the nanobody was isolated. Success of protein expression and secretion into the medium was evaluated using Western blot. In this research, we show that the suggested model system is applicable for production of the chosen nanobody.
Secondary keywords:	nanobody;Saccharomyces cerevisiae;secretion;Beljakovine;Protitelesa;Univerzitetna in visokošolska dela;
Type (COBISS):	Bachelor thesis/paper
Study programme:	1000371
Embargo end date (OpenAIRE):	1970-01-01
Thesis comment:	Univ. v Ljubljani, Fak. za kemijo in kemijsko tehnologijo, UNI Biokemija
Pages:	28 str.
ID:	19888195