CpG-free LacZnls

Synthetic LacZ reporter gene with SV40 nuclear localization signal in expression plasmid

The lacZ gene codes for the enzyme β-galactosidase which catalyzes the hydrolysis of the substrate X-Gal to produce a blue color that is easily visualized under a microscope.

The synthetic lacZ gene engineered by InvivoGen is entirely free of CpG motifs. LacZnls contains a nuclear localization signal of SV40 large T that allows the targeting of the chimeric protein to the nucleus.

The CpG-free LacZnls gene is provided in the expression plasmid pSelect-zeo which is selectable with Zeocin™ in both mammalian and E.coli cells.

Specifications

Gene: CpG-free LacZnls

Description: Synthetic β-Galactosidase with the SV40 nuclear localization signal

Backbone: pSELECT-zeo

Selection: Zeocin™

289 CpG in the native gene

Contents

• 20 µg of lyophilized DNA
• 1 ml of Zeocin™ (100 mg/ml)

Product is shipped at room temperature

Description

PLASMID FEATUES
First expression cassette
• hEF1-htLV prom is a composite promoter comprising the Elongation Factor-1alpha (EF-1α) core promoter [1 ]and the R segment and part of the U5 sequence (R-U5’) of the Human T-Cell Leukemia Virus (HTLV) Type 1 Long Terminal Repeat [2]. The EF-1α promoter exhibits a strong activity and yields long lasting expression of a transgene in vivo. The R-U5’ has been coupled to the EF-1α core promoter to enhance stability of RNA.
• LacZ-∆CpG NLS: The E. coli lacZ gene codes for the enzyme β-galactosidase which catalyzes the hydrolysis of the substrate X-Gal to produce a blue color that is easily visualized under a microscope. A nuclear localization signal of SV40 large T has been inserted in the 5’ end of the lacZ gene to allow the targeting of the chimeric protein to the nucleus. To reduce the immunogenicity of this bacterial gene, InvivoGen has engineered a synthetic lacZnls gene that is entirely free of CpG motifs, whereas the wild type lacZ gene contains 298 CpG dinucleotides.
• SV40 pAn: the Simian Virus 40 late polyadenylation signal enables efficient cleavage and polyadenylation reactions resulting in high levels of steady-state mRNA [3].
• ori: a minimal E. coli origin of replication to limit vector size, but with the same activity as the longer Ori.
Second expression cassette
• CMV enh/prom: The human cytomegalovirus immediate-early gene 1 promoter/enhancer was originally isolated from the Towne strain and was found to be stronger than any other viral promoters.
• EM7 is a bacterial promoter that enables the constitutive expression of the antibiotic resistance gene in E. coli.
• Zeo: Resistance to Zeocin™ is conferred by the Sh ble gene from Streptoalloteichus hindustanus The Sh ble gene is driven by the CMV enhancer/promoter in tandem with the bacterial EM7 promoter allowing selection in both mammalian cells and E. coli.
• ßGlo pAn: The human beta-globin 3’UTR and polyadenylation sequence allows efficient arrest of the transgene transcription [4].

1. Kim, D.W. et al. (1990). Gene 2: 217-223.
2. Takebe, Y. et al. (1988). Mol. Cell Biol. 1: 466-472.
3. Carswell, S. & Alwine, J.C. (1989). Mol. Cell Biol. 10: 4248-4258.
4. Yu J & Russell JE. (2001). Mol Cell Biol, 21(17):5879-88.