Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli

Kübra Özkul; Ebru Yılmaz; Aslıhan Kurt Kızıldoğan

doi:10.53433/yyufbed.1633412

TR EN

Fosfinotrisin N-asetiltransferazı Kodlayan Siyanobakteriyel Genlerin E. coli’de Klonlanması ve Heterolog İfadesi

Öz

Fosfinotrisin N-asetiltransferaz (Pat), fosfinotrisin (PPT) içeren herbisitlere direnç sağlamak için bitkilerde aktarılan bakteriyel bir enzimdir. Pat proteininin transgenik bitkilerde ifade edilmesi, glufosinat herbisitine karşı tolerans sağlar. Siyanobakterilerdeki bu enzim dizi analizi ile tanımlanmış olmasına rağmen, aktiviteleri henüz belirlenmemiştir. Bu çalışmada, Anabaena sp. PCC7120 ve Synechocystis sp. PCC6803’de Pat enzimini kodladığı varsayılan sırasıyla alr ve sll genleri His-Taq pET28a+ ekspesyon vektörüne klonlandı. İlgili genleri içeren rekombinant vektörler proteaz içermeyen E. coli BL21 hücrelerine transfer edildi. Alr4468 ve Sll1647 polipeptitleri 1 mM IPTG ile indüklenmiş rekombinant E. coli BL21 kültürlerinde başarıyla ifade edildi. En yüksek in vitro Pat aktiviteleri IPTG ile indüklenmiş kaba hücre ekstraktları kullanılarak yapılan enzim deneylerinde elde edilmiş ve kontrollerden sırasıyla 3.47 (rAlr4468 için) ve 2.53 (rSll1647 için) kat daha yüksek bulunmuştur. Sonuç olarak, yüksek yapılı bitkilerle benzer fizyolojik özelliklere sahip olan siyanobakterilerin alr4468 ve sll1647 genleri, gelecekteki çalışmalarda herbisite dirençli yerli transgenik bitkilerin üretimi için potansiyel kaynaklar olarak kullanılabilirler.

Anahtar Kelimeler

bar geni, Fosfinotrisin N-asetiltransferaz, Herbisit direnci, Siyanobakteriyel pat geni

Proje Numarası

PYO.ZRT.1902.16.001

Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli

Öz

Phosphinothricin N-acetyltransferase (Pat) is a bacterial enzyme that is introduced into plants to confer resistance to herbicides containing phosphinothricin (PPT). The expression of the Pat protein in transgenic plants allows them to tolerate the herbicide glufosinate. Although the enzyme has been identified in cyanobacteria through sequence analysis, its activities have not yet been thoroughly investigated. In this study, we cloned the putative Pat enzyme-encoding genes alr4468 and sll1647 from Anabaena sp. PCC7120 and Synechocystis sp. PCC6803, respectively, into the His-Tagged pET28a+ expression vector. The recombinant vectors containing these genes were introduced into protease-free E. coli BL21 cells. We successfully expressed the Alr4468 and Sll1647 polypeptides in the recombinant E. coli BL21 cultures that were induced with 1 mM IPTG. The highest in vitro Pat activities were measured using enzyme assays from IPTG-induced cell crude extracts, showing increases of 3.47 times for rAlr4468 and 2.53 times for rSll1647 compared to controls. As a result, the alr4468 and sll1647 genes from cyanobacteria, which exhibit similar physiological properties to those of high-structured plants, may serve as potential sources for developing herbicide-resistant transgenic plants in future research.

Anahtar Kelimeler

bar gene, Cyanobacterial pat gene, Herbicide resistance, Phosphinothricin N-acetyltransferase

Destekleyen Kurum

Ondokuz Mayıs University

Proje Numarası

PYO.ZRT.1902.16.001

Kaynakça

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Ayrıntılar

Birincil Dil

İngilizce

Konular

Mikrobiyal Genetik, Tarımda Enzim ve Mikrobiyal Biyoteknoloji

Bölüm

Araştırma Makalesi

Yazarlar

Kübra Özkul ^*
0000-0001-8958-4185
Türkiye

Ebru Yılmaz
0009-0002-8728-1919
Türkiye

Aslıhan Kurt Kızıldoğan
0000-0002-9323-0993
Türkiye

Yayımlanma Tarihi

31 Ağustos 2025

Gönderilme Tarihi

6 Şubat 2025

Kabul Tarihi

28 Mayıs 2025

Yayımlandığı Sayı

Yıl 2025 Cilt: 30 Sayı: 2

DOI

https://doi.org/10.53433/yyufbed.1633412

IZ

https://izlik.org/JA47ZR58SM

APA

Özkul, K., Yılmaz, E., & Kurt Kızıldoğan, A. (2025). Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli. Yüzüncü Yıl Üniversitesi Fen Bilimleri Enstitüsü Dergisi, 30(2), 815-829. https://doi.org/10.53433/yyufbed.1633412

AMA

1.Özkul K, Yılmaz E, Kurt Kızıldoğan A. Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli. YYUFBED. 2025;30(2):815-829. doi:10.53433/yyufbed.1633412

Chicago

Özkul, Kübra, Ebru Yılmaz, ve Aslıhan Kurt Kızıldoğan. 2025. “Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli”. Yüzüncü Yıl Üniversitesi Fen Bilimleri Enstitüsü Dergisi 30 (2): 815-29. https://doi.org/10.53433/yyufbed.1633412.

EndNote

Özkul K, Yılmaz E, Kurt Kızıldoğan A (01 Ağustos 2025) Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli. Yüzüncü Yıl Üniversitesi Fen Bilimleri Enstitüsü Dergisi 30 2 815–829.

IEEE

[1]K. Özkul, E. Yılmaz, ve A. Kurt Kızıldoğan, “Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli”, YYUFBED, c. 30, sy 2, ss. 815–829, Ağu. 2025, doi: 10.53433/yyufbed.1633412.

ISNAD

Özkul, Kübra - Yılmaz, Ebru - Kurt Kızıldoğan, Aslıhan. “Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli”. Yüzüncü Yıl Üniversitesi Fen Bilimleri Enstitüsü Dergisi 30/2 (01 Ağustos 2025): 815-829. https://doi.org/10.53433/yyufbed.1633412.

JAMA

1.Özkul K, Yılmaz E, Kurt Kızıldoğan A. Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli. YYUFBED. 2025;30:815–829.

MLA

Özkul, Kübra, vd. “Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli”. Yüzüncü Yıl Üniversitesi Fen Bilimleri Enstitüsü Dergisi, c. 30, sy 2, Ağustos 2025, ss. 815-29, doi:10.53433/yyufbed.1633412.

Vancouver

1.Kübra Özkul, Ebru Yılmaz, Aslıhan Kurt Kızıldoğan. Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli. YYUFBED. 01 Ağustos 2025;30(2):815-29. doi:10.53433/yyufbed.1633412

Fosfinotrisin N-asetiltransferazı Kodlayan Siyanobakteriyel Genlerin E. coli’de Klonlanması ve Heterolog İfadesi

Öz

Anahtar Kelimeler

Proje Numarası

Cloning and Heterologous Expression of Cyanobacterial Genes Encoding Phosphinothricin N- acetyltransferase in E. coli

Öz

Anahtar Kelimeler

Destekleyen Kurum

Proje Numarası

Kaynakça

Ayrıntılar

Birincil Dil

Konular

Bölüm

Yazarlar

Yayımlanma Tarihi

Gönderilme Tarihi

Kabul Tarihi

Yayımlandığı Sayı

DOI

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