Effect of storage time and temperatures on growth rate of Eschericia coli BL21 containing JSU construct for Jembrana disease candidate vaccine

Abstract

The viability of Eschericia coli BL21 (DE3) bearing JSU (Jembrana Surface Unit) recombinant protein depend on several factors, such as temperatures and time of storage. JSU act as a vaccine candidate for the Jembrana disease in Bali cattle, therefore, it is important to keep the quality and quantity the yield of JSU recombinant protein, the E. coli need to maintenance. The study was designed to examine the viability of Escherichia coli at different temperatures and time of cultivation. The E. coli BL21 bearing JSU was cultured in LB broth (+Amph 100µg mL−1) overnight and aliquot it in 1.5mL sterile tube (3 tube every temperatures). The tubes were incubated in three storage temperatures (Room Temperatures/RT, 4°C and -20°C. The viabilities of the cells were observed in 24, 48 and 120 hours by measuring the Optical density by spectrophotometer, and then inoculated it in LB plate (+Amp 100µg mL−1). The result showed that statistically, there were significant effect (P<0.05) in temperature -20°C at 24h and 120h observation on viability of E. coli BL21. The viability of E. coli at -20°C and storage time observed at 120h are more stabile, compared the other temperatures (RT and 4°C). There was no significant effect on both temperatures RT and 4°C in OD600 at 24 and 48h and the growth was decreased until 120h observation. The observation at 24h with 4°C cultured has a higher OD600 (1.098) followed by RT (1.070) and -20°C (0.773). The viability of E. coli was decreased at 48h with RT (1.011) and 4°C (1.0611), meanwhile, at -20°C the cells were increased (0.958). Meanwhile, the observation at 120h, the cells was more stabile at -20°C (0.961) compared with cultures from 4°C (0.981) and RT (0.719). Based on the OD600, the viability at the -20°C intended more stabile compared to RT and 4°C, but in the 24h the growth at -20°C are very low because no protectant added in the culture, it will shock the cells and dead. This finding showed that viability of E. coli BL21 bearing JSU recombinant protein better to keep in -20°C for longer period storage time with an addition of protectant agent in the culture media to keep the cells alive and more stabile.