Cloning and expression of SARS-COV-2 spike gene in bacteria and yeast
Küçük Resim Yok
Tarih
2023
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
İnönü Üniversitesi
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
Şiddetli Akut Solunum Sendromu Koronavirüs 2 (SARS-CoV-2), 2019 yılında Çin'in Wuhan şehrinde ortaya çıkan, koronavirüs hastalığına (COVID-19) neden olan, zarflı, pozitif anlamlı, tek sarmallı bir RNA virüsüdür. Virüs partikülleri bir RNA (mRNA), yapısal proteinler, yapısal olmayan proteinler (NSP'ler) ve yardımcı proteinler içerir. Virüs, konakçı hücrelerin yüzeyindeki reseptörler aracılığıyla hücreye girdikten sonra 4 yapısal protein; Spike, Membran, Nükleokapsid, Zarf (S, M, N, E), yaklaşık 20 yapısal olmayan protein (NSP'ler) ve bir düzine yardımcı protein (AP'ler) yapılır. Virüsün girişi, partikül oluşması ve bağışıklıktan kaçması için yapısal proteinler gerekliyken, NSP'ler ve AP'ler transkripsiyon, replikasyon ve konak hücre kontrolünde önemlidir. Virüsün en büyük protein olan Spike glikoproteini viral bağlanma ve konakçıya girişe aracılık ettiğinden aşı geliştirme, monoklonal antikor tedavileri ve antijen bazlı veya PCR tanı testleri için önemli bir hedef olmuştur. Bu çalışmada Spike geni hem bakteri hem de mayada replikasyon ve ekspresyon yapabilen bir mekik vektörüne klonlanmış ve proteinin ifade profili araştırılmıştır. Çalışmamız hem bakteri hem de mayanın Spike proteinini ifade ettiğini ve rekombinant mekik vektörünün her iki hücre tipinde de kararlı bir şekilde muhafaza edildiğini gösterdi. Anahtar Kelimeler: SARS-CoV-2, COVID-19, Koronavirüsler, Spike proteini, Mekik vektörler, Escherichia coli, Sacharomyces cerevisiae.
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is an enveloped, positive-sense, single-stranded RNA virus that causes coronavirus disease (COVID-19) that emerged in 2019 in Wuhan city of China. Virus particles contain an RNA (mRNA), structural proteins, non-structural proteins (NSPs), and accessory proteins. After the virus enters the cell via receptors on the surface of host cells, 4 structural proteins; Spike, Membrane, Nucleocapsid, Envelope (S, M, N, E), about 20 non-structural proteins (NSPs), and a dozen accessory proteins (APs) are made. While structural proteins are required for virus entry, assembly, and immune evasion, NSPs and APs are important in transcription, replication, and host cell control. Since the Spike glycoprotein, the largest protein of the virus, mediates viral attachment and entry into the host, it has been an important target for vaccine development, monoclonal antibody therapies, and antigen-based or PCR diagnostic tests. In this study, the Spike gene was cloned into a shuttle vector that can replicate and express in both bacteria and yeast, and the expression profile of the protein was investigated. Our study showed that both bacteria and yeast expressed the Spike protein and the recombinant shuttle vector was stably maintained in both cell types. Keywords: SARS-CoV-2, COVID-19, Coronaviruses, Spike protein, Shuttle vectors, Escherichia coli, Saccharomyces cerevisiae.
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is an enveloped, positive-sense, single-stranded RNA virus that causes coronavirus disease (COVID-19) that emerged in 2019 in Wuhan city of China. Virus particles contain an RNA (mRNA), structural proteins, non-structural proteins (NSPs), and accessory proteins. After the virus enters the cell via receptors on the surface of host cells, 4 structural proteins; Spike, Membrane, Nucleocapsid, Envelope (S, M, N, E), about 20 non-structural proteins (NSPs), and a dozen accessory proteins (APs) are made. While structural proteins are required for virus entry, assembly, and immune evasion, NSPs and APs are important in transcription, replication, and host cell control. Since the Spike glycoprotein, the largest protein of the virus, mediates viral attachment and entry into the host, it has been an important target for vaccine development, monoclonal antibody therapies, and antigen-based or PCR diagnostic tests. In this study, the Spike gene was cloned into a shuttle vector that can replicate and express in both bacteria and yeast, and the expression profile of the protein was investigated. Our study showed that both bacteria and yeast expressed the Spike protein and the recombinant shuttle vector was stably maintained in both cell types. Keywords: SARS-CoV-2, COVID-19, Coronaviruses, Spike protein, Shuttle vectors, Escherichia coli, Saccharomyces cerevisiae.
Açıklama
Anahtar Kelimeler
Biyomühendislik, Bioengineering, Mikrobiyoloji
