Hudson M C, Curtiss R
Department of Biology, Washington University, St. Louis, Missouri 63130.
Infect Immun. 1990 Feb;58(2):464-70. doi: 10.1128/iai.58.2.464-470.1990.
Studies were initiated to investigate the regulation of Streptococcus mutans genes which are believed to be important to virulence. Operon fusions were constructed between S. mutans gene regulatory regions and a promoterless chloramphenicol acetyltransferase gene (cat) found on the plasmid pMH109. Specifically, fusions were generated between cat and the S. mutans genes encoding fructosyltransferase (ftf) and the glucosyltransferase B/C (gtfB/C) operon. Constructs were confirmed by restriction enzyme analysis, and the fusions were subcloned into the integration vehicle pVA891. Following generation of multimeric DNA, recombinant plasmids were introduced into the s. mutans genome by Campbell-type insertion, resulting in single-copy operon fusions. Chloramphenicol acetyltransferase specific activities were used to monitor the expression of the S. mutans gtfB/C operon and ftf determinants. The expression of these genes is increased by the presence of sucrose and is followed by a rapid decline in expression over time. Additionally, expression of the gtfB/C operon is increased in S. mutans cells bound to artificial tooth pellicles.
已开展研究以调查变形链球菌中被认为对毒力很重要的基因的调控情况。在变形链球菌基因调控区域与质粒pMH109上发现的无启动子氯霉素乙酰转移酶基因(cat)之间构建了操纵子融合体。具体而言,在cat与编码果糖基转移酶(ftf)的变形链球菌基因以及葡糖基转移酶B/C(gtfB/C)操纵子之间产生了融合体。通过限制性酶切分析确认构建体,然后将融合体亚克隆到整合载体pVA891中。在产生多聚体DNA后,通过坎贝尔型插入将重组质粒引入变形链球菌基因组,从而产生单拷贝操纵子融合体。氯霉素乙酰转移酶的特异性活性用于监测变形链球菌gtfB/C操纵子和ftf决定簇的表达。这些基因的表达在蔗糖存在时增加,随后随着时间推移表达迅速下降。此外,在与人工牙菌斑结合的变形链球菌细胞中,gtfB/C操纵子的表达增加。