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All Journal Buletin Veteriner Udayana
I Gusti Ngurah Kade Mahardika
Biomedical and Veterinary Molecular Biology Laboratory, Faculty of Veterinary Medicine, Udayana University, Jl. Raya Sesetan, Gg. Markisa No. 6, Denpasar, Bali, Indonesia
Immunology & microbiology Public Health Veterinary

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THE EFFECT OF CONCENTRATION AND TIME VARIATION OF THE LACTOSE AND SKIM MILK AS INDUCER ON THE EXPRESSION OF RECOMBINANT PROTEIN BOVINE LACTOFERRIN CARBOXY-TERMINUS (BLF CAR) Febriyan Muji Hartini; I Gusti Ngurah Kade Mahardika; I Wayan Masa Tenaya
Buletin Veteriner Udayana Bul. Vet. Udayana. April 2026 Vol. 18 No. 2
Publisher : Fakultas Kedokteran Hewan Universitas Udayana

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24843/bulvet.2026.v18.i02.p04

Abstract

Recombinant bovine lactoferrin (bLf) production has broad potential applications in animal health, particularly as a feed additive and immunomodulatory agent, thereby requiring an efficient and sustainable production system. Lac operon–based expression systems commonly employ isopropyl β-D-1-thiogalactopyranoside (IPTG) as an inducer; however, its high cost and potential to induce metabolic stress limit its applicability for large-scale production. This study aimed to evaluate lactose and skim milk as alternative inducers for recombinant bLf expression. A synthetic bLf CAR gene was cloned into the pGEX-2TK plasmid to generate a GST-bLf CAR fusion protein and expressed under a lac operon–based system. Induction was performed using lactose and skim milk at concentrations of 1%, 2.5%, and 5%, with induction times of 5, 24, and 48 hours, while 0.1 mM IPTG served as the reference inducer. The experimental design followed a randomized block design. Protein expression was analyzed using dot blot, and signal intensity was quantified using ImageJ software. Statistical analysis was conducted using two-way ANOVA followed by Tukey ’s post hoc test. The results demonstrated that inducer type, concentration, and induction time significantly affected the expression intensity of recombinant GST-bLf CAR protein (p < 0.01). Skim milk at a concentration of 2.5% with a 5 hour induction period produced the highest expression level and differed significantly from other treatments. These findings indicate that skim milk is a promising, economical, and practical alternative inducer for recombinant bLf production.
THE EFFECT OF CONCENTRATION AND TIME VARIATION OF THE LACTOSE AND SKIM MILK ON THE EXPRESSION OF RECOMBINANT PROTEIN GST-A276R AFRICAN SWINE FEVER VIRUS Harry Prasetyo Triwijayanto; I Gusti Ngurah Kade Mahardika; I Nyoman Suartha
Buletin Veteriner Udayana Bul. Vet. Udayana. April 2026 Vol. 18 No. 2
Publisher : Fakultas Kedokteran Hewan Universitas Udayana

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24843/bulvet.2026.v18.i02.p07

Abstract

Recombinant protein expression in Escherichia coli (E. coli) commonly utilizes isopropyl β-D-1-thiogalactopyranoside (IPTG) as an inducer. However, IPTG causes toxicity, metabolic instability, and high cost. Lactose and skim milk provide safer and more economical alternatives and can induce the lac promoter in a more controlled manner. This study aimed to evaluate the effectiveness of lactose and skim milk as alternative inductors for expressing the GST-A276R fusion protein, a candidate subunit vaccine for African Swine Fever (ASF). The A276R gene from ASF virus was inserted into the pGEX-2TK plasmid to produce this fusion protein. Different inducer concentrations (1%, 2.5%, and 5%) and induction times (5, 24, and 48 hours) were evaluated in E. coli BL21 cultures using a randomized block design. Intensity of protein expression was analyzed by the dot blot method using anti-GST antibodies, and signal intensity was measured with ImageJ software. The results showed that the type of inducer had a significant effect on the expression level of GST-A276R protein (p < 0.05). The combination of 2.5% skim milk and 48 hours of skim milk induced expression was the most optimal setup for GST-A276R protein expression. In addition to inducer type, inducer concentration and induction time also had significant effects on protein expression (p < 0.05). Based on these findings, it is concluded that skim milk serves as an effective alternative inducer for GST-A276R protein production. Further analysis is required to test the purity and biological activity of the GST-A276R protein expressed under these optimal conditions.
Co-Authors Febriyan Muji Hartini Harry Prasetyo Triwijayanto I Nyoman Suartha I Wayan Masa Tenaya