pGlow TOPO™ TA Expression Kit

We do not recommend storing competent E. coli strains in liquid nitrogen as the extreme temperature can be harmful to the cells. Also, the plastic storage vials are not intended to withstand the extreme temperature and may crack or break.

We recommend storing our competent E. coli strains at -80°C. Storage at warmer temperatures, even for a brief period of time, will significantly decrease transformation efficiency.

pBlue-TOPO contains a cryptic prokaryotic promoter upstream of the lacZ reporter gene, due to which E. coli transformants may appear to be light blue when screened on plates containing X-Gal. Hence, we do not recommend using pBlue-TOPO to evaluate promoter function in E. coli. However, pGlow-TOPO can be used for these studies. Note that background expression of beta-galactosidase from pBlue-TOPO does not occur in mammalian cells.

Yes, we do offer the pBlue-TOPO and pGlow-TOPO vectors that facilitate cloning of the DNA sequence of interest directly upstream of either the b-galactosidase or Cycle 3 GFP gene, respectively.
pBlue-TOPO is ideal for functional analysis of promoters with low transcriptional activity, since assays for beta-galactosidase are easy to perform and are quantitative at very low levels of expression. pGlow-TOPO is ideal for non-invasive analysis of promoter elements within intact, living cells. The fluorescent property of Cycle 3 GFP allows in vivo detection in virtually any cell type or species using microscopy with wild-type GFP filter sets or by fluorescence-activated cell sorting methods.

We recommend looking for GFP fluorescence before staining for beta-galactosidase. This is because the beta-galactosidase staining process produces a very high autofluorescence that will interfere with detection of GFP fluorescence.