|
Transfer and expression of exendin-4 fused to cholera toxin B subunit (CTB) in lettuce
|
Ebrahim Moradi   , Mohammad Naji , Bahman Bahramnejad *  , Hemn Salehi |
| Department of Plant production and Genetics, Faculty of Agriculture, University of Kurdistan, Sanandaj, Iran , b.bahramnejad@uok.ac.ir |
|
|
Abstract: (1118 Views) |
Exendin-4 (EX4) is a protein similar to glucan-like peptide (GLP-1) and has a longer half-life and greater effect in inducing insulin secretion. The fusion of EX4 to the non-toxic cholera subunit (CTB) increases its efficiency through absorption in the digestive system. In this study, CTB-EX4 gene was introduced into lettuce by Agrobacterium tumefaciens using vector pBI121. The results showed that 20.25% of the inoculated explants were successfully transformed and regenerated (T0 generation). To create T2 plants, seeds of T1 were cultivated, and 42% of the cultivated seeds germinated. T1 plants were analyzed using PCR test. The mRNA expression of the CTB-EX4 gene was examined through RT-PCR and Real-time PCR, while the protein expression was assessed using ELISA and Western Blot tests. The number of transferred copies to T1 plants was determined using Real-time PCR. This estimation was performed using both the absolute method and the standard curve, as well as the relative method, with Acitn gene as a reference gene. The results showed that 60% of the T1 plants were transgenic and the CTB-EX4 gene was expressed at the mRNA and protein levels. The results showed that the process of transferring the CTB-EX4 gene to the lettuce plant was successful and the lettuce plant was able to express the desired gene at the RNA and protein levels. Considering the subsequent processes, including the purification and evaluation of the biological activity of the produced protein, plants can be used as a suitable bioreactor for the production of recombinant proteins. |
|
| Keywords: Agrobacterium tumefaciens, Exendin, gene expression, pBI121 vector, Recombinant protein. |
|
|
Full-Text [PDF 1348 kb]
(298 Downloads)
|
Type of Study: Research |
Subject:
Plant
Received: 2023/07/2 | Accepted: 2023/08/29 | Published: 2024/03/14
|
|
|
|
|
|
|
| Add your comments about this article |
|
|
|
