Presently we offer vectors for targeting your protein to 5 different subcellular localizations. Targeting a protein or enzyme to a different environment can produce strongly different phenotypes. Enzymes may find completely new or higher levels of substrates. Alternatively, proteins may be less degraded due to the absence of specific proteases and accumulate much higher levels. The provided targeting options have been either directly tested at Plant Research International or have been taken from the available literature on the subject.

ImpactVector 1.1 Cytoplasmic expression or expression with native signals
By default genes or gene fragments without pre-pro targeting peptides will express in the cytoplasm and this basic vector will help to achieve just that. It does provide the option of adding a tag to your protein of interest. The vector is also your choice if you want to express your gene with its own native targeting information.

ImpactVector 1.2 Secreted expression
Secreted expression is important for proper formation of disulphide bonds, glycosylation, etc., and is the first step in the targeting of proteins to the ER, Golgi, transport vesicles, the vacuole, peroxisomes etc. The default pathway is secretion out of the cell into the cell wall and apoplast. Various sorting and retention signals determine the actual protein destination (see ImpactVector1.3). The vector can be used to change the targeting of the mature part of a protein to a secreted destination.The signal peptide used in this construct was derived from sea anemone equistatin (Outchkourov et al. 2003).

ImpactVector 1.3 Endoplasmic reticulum expression
By adding the well known KDEL retention or sorting signal to ImpactVector1.2 we created vector RbcS1.3. With this vector proteins are retained in the ER and do not go downstream into Golgi. ER retention has a practical advantage in that it has been reported to improve expression levels 5-fold or more. The main reason for this appears to be that the ER contains lower concentrations and/or different proteases responsible for post translational degradation of expressed proteins (Outchkourov et al. Sept 2003 Plant Physiology in press)

ImpactVector 1.4 Chloroplast expression
Targeting to the chloroplast brings a protein into a completely different cellular environment capable of different metabolic and catabolic functions. ImpactVector1.4 vector uses the secretion signal of the Chrysanthemum morifolium small subunit protein to deliver your protein into the chloroplast stroma (cytoplasmic) environment shuttling across a double membrane system. The protein is fused to the first 11 amino acids of the mature rubisco protein in order to allow proper processing of the signal peptide (Wong et al., Plant Molecular Biology 20: 81-93 (1992)). The signal peptide contains a natural intron from the RbcS gene.

ImpactVector 1.5 Mitochondrial expression
Targeting to mitochondria brings a protein to a completely different subcellular environment capable of different metabolic and catabolic functions. The ImpactVector1.5 vector uses the yeast CoxIV secretion signal, which was shown to deliver proteins in the mitochondrial matrix. The protein is fused to the first 4 amino acids of the yeast CoxIV protein in order to allow proper processing of the signal peptide (Kohler et al. Plant J 11: 613-621 (1997)).