A bi-functional reporter plasmid for the simultaneous transient expression assay of two herpes simplex virus promoters.
A bi-functional reporter plasmid for the simultaneous transient expression assay of two herpes simplex virus promoters.
复制标题
用于同时瞬时表达测定两种单纯疱疹病毒启动子的双功能报告质粒。
DOI：
10.1007/bf00125686
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发表时间：
1987
期刊：
影响因子：
1.6
通讯作者：
Wagner,EK
中科院分区：
文献类型：
--
作者：
Flanagan,WM;Wagner,EK
我们构建了一个含有氯霉素乙酰转移酶（CAT）和β-半乳糖苷酶（β-gal）两个报告基因的瞬时表达载体（pCAL），用于研究单纯疱疹病毒1型（HSV-1）启动子在哺乳动物细胞中的活性。该构建体被设计用于分析来自两个不同启动子的同时表达。为了测试该载体的实用性，我们使用了三个HSV-1启动子，这些启动子先前已经由本实验室的工作人员进行了表征。两个是早期（β）启动子，用于碱性核酸外切酶和脱氧尿苷三磷酸核苷酸水解酶;第三个启动子控制主要衣壳蛋白转录物并且是晚期（βγ）。将两种不同动力学类型的启动子以不同的方向连接到pCAL中并转染到兔皮肤成纤维细胞中。然后用低多样性的HSV-1超感染转染的细胞; 18小时后，我们观察到两种标记基因在各自启动子的控制下同时表达。这种瞬时表达报告载体的有用性进行了讨论。
We have constructed a transient expression vector (pCAL) containing two reporter genes, chloramphenicol acetyltransferase (CAT) and β-galactosidase (β-gal) for use in studying herpes simplex virus type 1 (HSV-1) promoter activity in mammalian cells. The construct was designed to be useful in analyzing the simultaneous expression from two different promoters. To test the utility of the vector, we used three HSV-1 promoters that had been characterized previously by workers in this laboratory. Two are early (β) promoters, for alkaline exonuclease and deoxyuridine triphosphate nucleotidohydrolase; the third promoter controls the major capsid protein transcript and is late (βγ). The two different kinetic classes of promoters were ligated in a divergent orientation into pCAL and transfected into rabbit skin fibroblast. Transfected cells were then superinfected with low multiplicities of HSV-1; 18 hr later, we observed the simultaneous expression of both marker genes under control of the respective promoters. The usefulness of such a transient expression reporter vector is discussed.