Prasad R, Niederberger P, Hütter R
Mikrobiologisches Institut, Eidgenössische Technische Hochschule, Zürich, Switzerland.
Yeast. 1987 Jun;3(2):95-105. doi: 10.1002/yea.320030206.
Plasmid pME559, carrying all five yeast TRP genes, was constructed. This plasmid is a yeast/Escherichia coli shuttle vector based on pBR322 and 2 micron-DNA sequences derived from plasmid pJDB207. We studied in yeast (i) the stability of the plasmid under selective and non-selective conditions, (ii) expression of all five TRP genes and (iii) tryptophan accumulation in yeast transformants. These studies were conducted in comparison with an earlier construction, pME554, which differs from plasmid pME559 in the expression of the TRP1 gene and which carries the TRP2 wild type instead of the TRP2fbr mutant allele. For stable maintenance of the plasmids in yeast a selection was necessary. Plasmid pME559 displayed normal expression of all TRP genes, and enzyme levels on average 23-fold higher than in the wild type strain were found. In comparison, the maximal tryptophan flux observed in such a plasmid-carrying strain was about ten-fold higher than the maximal flux capacity in the wild type strain.
构建了携带所有五个酵母色氨酸（TRP）基因的质粒pME559。该质粒是基于pBR322和源自质粒pJDB207的2微米-DNA序列的酵母/大肠杆菌穿梭载体。我们在酵母中研究了：（i）该质粒在选择性和非选择性条件下的稳定性；（ii）所有五个TRP基因的表达；（iii）酵母转化体中的色氨酸积累。这些研究是与早期构建的pME554进行比较，pME554与质粒pME559的不同之处在于TRP1基因的表达，并且携带TRP2野生型而非TRP2fbr突变等位基因。为了在酵母中稳定维持质粒，选择是必要的。质粒pME559显示出所有TRP基因的正常表达，并且发现酶水平平均比野生型菌株高23倍。相比之下，在这种携带质粒的菌株中观察到的最大色氨酸通量比野生型菌株中的最大通量能力高约十倍。
