Page 8 - O. Microbiology

[O. Microbiology-10]

Production of recombinant alcohol dehydrogenase from

transformed Escherichia coli

Minhye Kim¹, Eunju Im¹, Yeh-Jin Ahn¹˙*

¹Biotechnology, Sangmyung University, Seoul 03016, Korea

Producing recombinant proteins from Escherichia coli(E. coli) has been widely used in biotechnology. In this study,
we developed transgenic E. coli cell lines heterologously expressing the heat shock protein 70 of carrot (Daucus
carota L.). DcHsp70 gene was attached to the DnaK promoter and FRT cassette containing a kanamycin-resistant

gene by overlap PCR, and then the DnaK promoter – DcHsp70 – FRT construct was inserted into the yddE site of

the E. coli genome via Red/ET mediated homologous recombination. To examine the expression level of recombinant
proteins, we introduced a recombinant pET11a plasmid containing 6His tagged alcohol dehydrogenase (ADH) into
the transgenic cell lines. When the recombinant 6His-ADH was expressed by isopropyl β-D-1-thiogalactopyranoside

treatment, the transgenic cell lines showed up to 10 times higher production of his-tagged ADH mainly in soluble
forms, compared to unmodified E. coli control cell line. Our results suggest that transgenic cell lines expressing

DcHsp70 could be a useful host E. coli cell line for recombinant protein production.