Open AccessHeat shock transformation and expression of the plasmid containing cytolethal distending toxin of Campylobacter fetus subsp venerealis in Escherichia coli
Author(s) -
Nina Herlina,
Nova Dilla Yanthi,
Riyona Desvy Pratiwi,
Kartika Sari Dewi,
Fitri Setiyoningrum,
Dadang Priyoatmojo,
R D P Manggung
Publication year - 2021
Publication title -
iop conference series. earth and environmental science
Language(s) - English
Resource type - Journals
eISSN - 1755-1307
pISSN - 1755-1315
DOI - 10.1088/1755-1315/888/1/012021
Subject(s) - escherichia coli , lac operon , transformation (genetics) , plasmid , heat shock protein , microbiology and biotechnology , biology , toxin , cytolethal distending toxin , recombinant dna , protein subunit , campylobacter fetus , heat labile enterotoxin , enterotoxin , campylobacter , gene , bacteria , biochemistry , genetics , microbial toxins
The cytolethal distending toxins (cdt) is a multi-subunit toxin consisted of three subunit encoded cdtA, cdtB and cdtC. The cdt played an important role as a virulence factor of Campylobacter infection, including C. fetus subsp venerealis . The cdtA which responsible for binding the cdt to cell membrane, was cloned in plasmid expression and inserted into bacterial cells of Escherichia coli BL21(DE3). The research was conducted to evaluate the transformation using the heat shock method of a plasmid containing cdtA3 gene and the protein expression induced by various concentration of IPTG. Transformation was done using the heat shock method at 42 o C for 90 second. Evaluation of the transformation was observed on the presence of E. coli BL21(DE3) colonies on Luria Bertani agar containing Ampicillin antibiotic with 100 µg/mL dosage. The recombinant protein was expressed using IPTG-induction with various concentration (0.1mM, 0.25mM, 0.5mM, 0.75mM and 1 mM). The result showed that the transformation and IPTG-induction 0.1 mM produced higher concentration of protein than other concentration applied. The protein characterization was observed with SDS PAGE and cdtA3 protein was detected on 23,4 kDa.