Soluble Expression of Synthetic CSF3syn Gene Fused with Thioredoxin in Escherichia Coli BL21(DE3) Through Autoinduction Method and Purification  

Oleh:

Pratiwi, Riyona Desvy ; Fuad, Asrul Muhamad

Jenis: Article from Journal - ilmiah nasional - terakreditasi DIKTI
Dalam koleksi: Indonesian Journal of Pharmacy vol. 26 no. 02 (Apr. 2015), page 63-70.
Topik: hG-CSF; Thioredoxin; Autoinduction; IMAC; E.Coli
Fulltext: I03 v26 n2 p63 kelik2017.pdf (796.01KB)

Ketersediaan

Perpustakaan FK Nomor Panggil: I03.K Non-tandon: 1 (dapat dipinjam: 0) Tandon: tidak ada

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Isi artikel

A synthetic human gene of CSF3 (CSF3syn.Ec3), coding for hG-CSF was succesfully subcloned into pET32a(+) expression vector and fused with thioredoxin (Trx) at its N-terminal as fusion partner. The obtained fusion gene of Trx-CSF3syn within the recombinant plasmid pET32a(+)_CSF3yn.Ec3 was verified by PCR, plasmid restriction, and DNA sequencing analysis. In order to investigate the fusion gene expression, we transformed Escherichia coli BL21(DE3) as the host with the recombinant plasmid. The gene was succesfully expressed within the cytosol as fusion protein of Trx·tag, His·tag, S·tag, EK-site, and hG-CSF moieties. By the auto induction method, 49% of the protein was found in the soluble fraction and the other 51% was found in the insoluble fraction. The soluble fraction was subsequently purified by IMAC method (Ni-NTA) and characterized.

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