Genski elektroprenos DNA-cepiva proti antigenom virusa SARS-CoV-2 v mišje mioblaste C2C12

Reberšek, Eva (avtor), Čemažar, Maja (mentor), Jesenko, Tanja (komentor)

Posledice pandemije, ki je sledila pojavu novega koronavirusa SARS-CoV-2 leta 2019 na Kitajskem, se kažejo v preobremenjenosti zdravstvenih sistemov, globalni ekonomiji in geopolitiki ter spremenjenih socialno-ekonomskih razmerah. Razvoj učinkovitih cepiv je zato ključnega pomena. DNA-cepiva so zaradi enostavnosti za pripravo, cenovne ugodnosti, dolgotrajne obstojnosti in specifičnega imunskega odziva proti želenemu antigenu uspešna metoda imunizacije, primerna tudi za razvoj cepiv proti virusu SARS-CoV-2. Genski elektroprenos je učinkovita metoda za vnos plazmidne DNA v celice ali tkiva, zato smo jo v naši raziskavi uporabili za vnos komercialno dostopnih plazmidov z zapisom za antigene virusa SARS-CoV-2 in sicer za protein bodice (protein S) in nukleokapside (protein N). Študijo smo izvedli in vitro na celični liniji mišjih mioblastov C2C12, kot modelnih celicah za mišično tkivo. Oba gena s plazmidov sta se izražala na ravni mRNA, na ravni proteinov pa smo zaznali le protein N, tako v celičnem mediju kot v celičnem lizatu. Genski elektroprenos obeh plazmidnih vektorjev skupaj je vodil do izražanja obeh antigenov na ravni mRNA v enaki meri kot pri genskem elektroprenosu posameznih plazmidnih DNA. Na ravni proteinov smo po genskem elektroprenosu obeh plazmidnih vektorjev hkrati uspeli dokazati le protein N. Protein je torej sekretoren, kar je dobra iztočnica za njegovo uporabo v namen vakcinacije. Rezultati naše raziskave dajejo dobro podlago za nadaljevanje študije in vivo na mišjih modelih, na katerih bi lahko preučevali učinkovitost vnosa plazmidne DNA z zapisom za antigene virusa SARS-CoV-2 v mišico in spremljali specifičen imunski odziv.

• Jezik: Slovenski jezik
• Ključne besede: genski elektroprenos, DNA-cepivo, virus SARS-CoV-2, protein S, protein N
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: BF - Biotehniška fakulteta
• Leto izida: 2023
• PID: 20.500.12556/RUL-143723
• Datum objave v RUL: 11.01.2023

REBERŠEK, Eva, 2023, Genski elektroprenos DNA-cepiva  proti antigenom virusa SARS-CoV-2 v mišje mioblaste C2C12 [na spletu]. Magistrsko delo. [Dostopano 21 april 2025]. Pridobljeno s: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=slv&id=143723

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Gene electrotransfer of DNA vaccine against SARS-CoV-2 viral antigens in C2C12 murine mioblasts

Izvleček

The effects of the pandemic following the emergence of a new SARS-CoV-2 coronavirus in China in 2019 are visible in the strain on health care systems, global economy and geopolitics, and changing socio-economic conditions. The development of effective vaccines is therefore crucial. DNA vaccines are a successful immunisation method due to their ease of preparation, cost-effectiveness, long-term durability and specific immune response against the desired antigen and are also suitable for the development of SARS-CoV-2 vaccines. Gene electrotransfer is an efficient method for introducing plasmid DNA into cells or tissues, and in our study we used it to introduce commercially available plasmids with sequences for SARS-CoV-2 antigens, namely the spike protein (S protein) and the nucleocapsid (N protein). The study was performed in vitro on the murine myoblast cell line C2C12, as model cells for muscle tissue. Both plasmid genes were expressed at the mRNA level, but only the N protein was detected at the protein level, both in the cell medium and in the cell lysate. Gene electroporation of both plasmid vectors together lead to the expression of both antigens at the mRNA level to the same extent as after gene electroporation of the individual plasmid DNA. At the protein level, we were only able to demonstrate the N protein after gene transfer of both plasmid vectors at the same time. The results of our study form a good basis for further in vivo studies in mouse models to demonstrate the effectiveness of introducing plasmid DNA with a transcript for SARS-CoV-2 antigens into muscle tissue and to monitor the specific immune response.

• Ključne besede: gene electrotransfer, DNA vaccine, virus SARS-CoV-2, S protein, N protein