2013 한국미생물학회연합 국제학술대회, 2013, 02, 319─319

RNA interference-mediated baculovirus-replication interference for reduction of residual baculovirus in virus-like particle production

HanSaem Lee, Ho Yeon Lee, Hee-Dong Jung, You-Jin Kim, Jeongsun Yang, Kisoon Kim

Abstract

The baculovirus expression system has been widely used to produce large amounts of post-translation modified recombinant proteins and virus-like particles (VLPs) as vaccine candidates for the prevention of serious infectious diseases. However, during bioprocess of VLPs in insect cells, baculovirus replication and budding were occurred coincidently. In some cases, residual baculovirus contaminants are still remained in VLPs, though various purification processes, ion-exchange chromatography, ultracentrifugation, or gel filtration, have been applied. Both glycoprotein 64 (GP64) and ssDNA binding protein (DBP) play critical roles in the assembly of the budded virion. To reduce unexpected contamination originated from the baculovirus replication, the siRNAs targeting GP64 and DBP were designed to inhibit baculovirus assembly without the hindrance of foreign gene expression. Here, GP64 expression was suppressed by GP64-targeted siRNAs in immunoblot experiments, while the foreign GFP protein expression was not affected. Moreover, transfection of GP64- and DBP-targeted siRNAs reduced the level of baculovirus replication, compared to scramble siRNAs treatment. In conclusion, GP64-targeted siRNAs showed potential that RNAi system can be applicable to reduce baculovirus remnants in VLP products. Further investigation should be accomplished to establish stable insect cell lines expressing corresponding RNAi.

• 본 연구는 질병관리본부 연구개발과제(과제번호 2012-N47002-00) 연구비를 지원받아 수행되었습니다.
• This research was supported by a fund(code 2012-N47002-00) by Research of Korea Centers for Disease Control and Prevention.