B형 간염 바이러스 표면항원 유전자가 효모 GAL10, ADCI 혹은 GAL1-ADCI double promoter 조절하에 발현되도록 재조합 플라즈미드를 제조하였다. Double prornoter는 ADCI promoter DNA절편과 GAL1-GAL10 promoter DNA 절편을 결합시킴으로써 만들었다. 효모에서 세가지의 재조합 플라즈미드는 모두 HBsAg을 발현시켰다. GAL promoter가 유도되었을 때 ADCI promoter 보다 3-5 배 많은 HBsAg을 생산하였다. Double promoter 시스템에서 galactose에 의해 전사가 유도되는 것이 확인되었지만 HBsAg 생성량은 유도되기 전의 수준에서 크게 증가되지 않았다. Northern blot에 의해 RNA 분석 결과 윗쪽 GAL promoter 가 유도됨에 따라서 아랫쪽 ADCI promoter 조절하의 전사시작이 저해 되고 있음이 시사되었다.
We have constructed plasmids in which transcription of the gene encoding human hepatitis B virus surface antigen(HBsAg) is under the control of GAL10, ADCI or a double promoter consisting of GAL1 and ADCI. Construction of the double promoter was achieved by insertion of a 650 bp fragment of divergent GAL1-GAL10 promoter DNA in the upstream of ADCI promoter. By radioimmunoassay, it was shown that three recombinant plasmids were all capable of producing HBsAg in the host Saccharomyces cerevisiae strains. When induced by galactose, cells synthesized 3-5 fold more HBsAg under GAL promoter control than under ADCI promoter control in an identical plasmid. Production of HBsAg in a system utilizing the double promoter was not increased additively, although it appeared to retain galactose inducibility. Northern blot analysis indicated that induction of the upstream GAL1 promoter caused inhibition of transcription initiation at the downstream ADCI promoter.
