Gateway® Case12-mito entry clone

cat.# FP995

The vector sequence has been compiled using the information from sequence databases, published literature, and other sources, together with partial sequences obtained by Evrogen. This vector has not been completely sequenced.

Download vector information: Data sheet (pdf) Sequence (txt) Restriction map (pdf) Restriction table (pdf)

Product Cat.# Size Price Gateway® Case12-mito FP995 20 μg € 600 The price does not include delivery. The price varies in different countries. Please contact your local distributor for exact prices and delivery information. The product is not available in certain countries. Please contact Evrogen or your local distributor before ordering. Vector type Gateway® entry clone Reporter Case12 Reporter codon usage mammalian Promoter for Case12 NO Host cells prokaryotic Selection kanamycin Replication pUC ori Use Transfer of Case12-mito coding sequence into Gateway® destination vectors

Vector description

Gateway® Case12-mito entry clone is a vector containing coding sequence of mitochondria-targeted fluorescent sensor Case12 (see reporter description). Mitochondrial targeting sequence (MTS) is fused to the Case12 N-terminus. MTS was derived from the subunit VIII of human cytochrome C oxidase [Rizzuto et al., 1989; Rizzuto et al., 1995]. Case12-mito coding sequence is flanked by attL1 and attL2 sites allowing easy site-specific recombination. The Invitrogen Gateway® Technology provides a rapid and highly efficient way to transfer the Case12-mito gene into a number of Gateway® destination vectors for expression in different experimental systems.

The vector backbone contains pUC origin of replication and kanamycin resistance gene (Kan^r) for propagation and selection in E. coli.

LR site-specific recombination

Please refer to Invitrogen Gateway® Technology description for detailed instructions regarding LR site-specific recombination reaction. In general, to transfer Case12-mito gene into the destination vector you will need:

  - Purified plasmid DNA of Gateway® Case12-mito
  - A destination vector of choice
  - Invitrogen LR Clonase^TM II enzyme mix (Invitrogen Cat.# 11791-020)
  - Proteinase K solution (supplied with the LR Clonase^TM II enzyme mix)
  - TE-Buffer, pH 8.0 (10 mM Tris-HCl, pH 8.0, 1 mM EDTA)
  - Appropriate chemically competent E. coli host and growth media for expression
  - Appropriate selective plates.

Propagation in E. coli

Suitable host strains for propagation in E. coli include DH5alpha, HB101, XL1-Blue, and other general purpose strains. Plasmid incompatibility group is pMB1/ColE1. The vector confers resistance to kanamycin (30 μg/ml) to E. coli hosts. Copy number in E. coli is about 500.

Location of features

attL1 site: 14-113
Case12-mito: 120-1475
attL2 site: 1494-1593
Kanamycin resistance gene: 2811-3605
pUC origin of replication: 4190-4833