Ozeki K, Kanda A, Hamachi M, Nunokawa Y
General Research Laboratory, Ozeki Corporation, Hyogo, Japan.
Biosci Biotechnol Biochem. 1996 Mar;60(3):383-9. doi: 10.1271/bbb.60.383.
We used a plasmid carrying a sequence for autonomous maintenance in Aspergillus (AMA1) and the E. coli uidA gene as a reporter gene to search the A. oryzae and A. niger genomes for DNA fragments having strong promoter activity. Beta-glucuronidase (GUS)-producing A. oryzae transformants containing the No. 8AN derived from A. niger, or the No. 9AO derived from A. oryzae, were constitutive for the expression of the uidA gene when cultivated in the presence of a variety of carbon and nitrogen sources. When the GUS-producing transformants were grown in liquid culture, the No. 8AN showed an increase of approximately 3-fold in GUS activity compared to the amyB (alpha-amylase encoding gene) promoter. There was also a corresponding increase in the amount of GUS gene-specific mRNA. When these transformants were grown as rice-koji, the No. 8AN showed an increase of approximately 6-fold compared to the amyB promoter, and the amount of GUS protein produced also increased. These strong promoter regions might be applicable to the production of other heterologous proteins in Aspergillus species.
我们使用了一个携带在曲霉中自主维持序列（AMA1）的质粒，并将大肠杆菌uidA基因作为报告基因，在米曲霉和黑曲霉基因组中搜索具有强启动子活性的DNA片段。含有源自黑曲霉的8AN或源自米曲霉的9AO的产生β-葡萄糖醛酸酶（GUS）的米曲霉转化体，在多种碳源和氮源存在下培养时，uidA基因的表达是组成型的。当产生GUS的转化体在液体培养中生长时，与amyB（编码α-淀粉酶的基因）启动子相比，8AN的GUS活性增加了约3倍。GUS基因特异性mRNA的量也相应增加。当这些转化体作为米曲培养时，与amyB启动子相比，8AN增加了约6倍，并且产生的GUS蛋白量也增加了。这些强启动子区域可能适用于在曲霉属物种中生产其他异源蛋白。
