■ 基本信息
启动子: | CMV |
复制子: | pUC |
终止子: | bGH poly(A) signal |
质粒分类: | 哺乳系列质粒；哺乳表达质粒；pCMV系列质粒 |
质粒大小: | 6210bp |
原核抗性: | Amp |
真核抗性: | Zeo |
克隆菌株: | DH5a |
培养条件: | 37℃，5%CO2 |
表达宿主: | 293T等哺乳细胞 |
诱导方式: | 无需诱导 |
5'测序引物:
| CMV-F（CGCAAATGGGCGGTAGGCGTG）
|
3'测序引物: | BGH（TAGAAGGCACAGTCGAGG） |
■ 质粒属性
质粒宿主: | 哺乳细胞 |
质粒用途: | 蛋白表达 |
片段类型: | ORF
|
片段物种: | 空载体
|
原核抗性: | Amp |
真核抗性: | Zeo
|
荧光标记: | 绿色
|
■ 质粒简介
pTracer-CMV2的CMV启动子后有MCS供插入外源基因，hEF1α启动子调控Cycle 3 GFP-Zeocin fusion的表达，可以做到既有荧光蛋白来做转染指示，且荧光基因表达和外源基因表达又不会互相干扰，而且Zeocin也可以用来做稳定性细胞筛选。另外，由于EM-7启动子的存在，Cycle 3 GFP-Zeocin fusion在E.coli中也可以表达，可以用来做原核荧光定位。
pTracer-CMV2 is a 6.2 kb mammalian expression vector derived from pcDNA3.1 and designed for visual detection of transformed E. coli cells and transfected mammalian cells. The vector contains the following elements:
Cycle 3-GFP, an improved GFP (Green Fluorescent Protein) gene (Crameri et al.,1996), fused to the Zeocin™ resistance gene (see below) for convenient,noninvasive detection of transformed or transfected cells
• Human elongation factor 1-subunit promoter (hEF-1) for mammalian expression of the Cycle 3-GFP-Zeocin fusion (Goldman et al., 1996; Mizushima and Nagata, 1990)
• A synthetic bacterial promoter, EM-7, for expression of Cycle 3-GFP in E. coli
• Human cytomegalovirus immediate-early (CMV) promoter for high-level expression of your gene in a wide range of mammalian cells
• Zeocin™ resistance for stable selection in mammalian cell lines
• Ampicillin and Zeocin resistance genes for selection of transformants in E. coli
The Cycle 3-GFP gene used in this vector is described in Crameri et al., 1996. In this paper, the codon usage was optimized for expression in mammalian cells and three cycles of DNA shuffling were used to generate a mutant form of GFP that has the following characteristics:
• Excitation and emission maxima that are the same as wild-type GFP (395 nm and 478 nm for primary and secondary excitation, respectively, and 507 nm for emission)
• High solubility in E. coli for visual detection of transformed cells
• >40-fold increase in fluorescent yield over wild-type GFP
• The Cycle 3-GFP gene is fused to the Zeocin™ resistance marker to correlate GFP fluorescence with resistance to the antibiotic Zeocin.
Zeocin is a member of the bleomycin/phleomycin family of antibiotics isolated from Streptomyces. It shows strong toxicity against bacteria, fungi (including yeast), plants, and mammalian cell lines. It is particularly well-suited for selection of mammalian stable cell lines. Ordering information is provided below.
质粒只保证关键序列正确，不保证表达效果。
■ 质粒图谱
■ 质粒序列
质粒序列请下载：
ZK388pTracer-CMV2哺乳表达质粒.txt
