Expression of hepatitis B surface antigen in tobacco cell suspension cultures.
Hepatitis B virus ' s ' gene coding for surface antigen was cloned into plant transformation vectors pHER100 and pHBs100 with and without endoplasmic reticulum retention signal, respectively. Transformed tobacco cell lines were analyzed for the integration of the transgene by PCR and Southern blot hybridization. Expression levels as determined by ELISA showed maximum expression levels of 2 microg HBsAg gm(-1) fresh weight and 10 ng mL(-1) of spent medium in pHER100 transformed cells. Western blot analysis confirmed the presence of 24 kDa band specific to HBsAg in the transformed cells. HBsAg was expressed both as intracellular and secreted forms in pHER100 transformed cells. The buoyant density in CsCl of HBsAg derived from pHBs100 transformed tobacco cells was determined and found to be 1.095 g mL(-1). HBsAg obtained from transformed tobacco cells is similar to the human serum derived one in buoyant density properties. This is the first report on the secretion of HBsAg particles by plant cells into the cell culture medium.
Sunil Kumar
G B
GB
Plant Cell Culture Technology Section, Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400 085, India.
Ganapathi
T R
TR
Revathi
C J
CJ
Prasad
K S N
KS
Bapat
V A
VA
D015139:Blotting, Southern; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D002586:Cesium; D002712:Chlorides; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors / Q000235:genetics; D006514:Hepatitis B Surface Antigens / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification*; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D013535:Suspensions; D014026:Nicotiana / Q000166:cytology / Q000235:genetics*; D014170:Transformation, Genetic
D002712:Chlorides; D006514:Hepatitis B Surface Antigens; D013535:Suspensions; D002586:Cesium; C028019:cesium chloride