Motif trap: a rapid method to clone motifs that can target proteins to defined subcellular localisations.
We have developed a rapid procedure termed Motif Trap (MT) to identify protein motifs that are able to target proteins to a distinct subcellular localisation in eukaryotic cells. By expressing random DNA fragments fused to green fluorescent protein (GFP), individual cells with the GFP localisation of interest are readily isolated allowing for the expressed DNA fragments to be cloned by RT-PCR. These can then be used to identify the corresponding full-length cDNAs. Using MT, we have identified patterns of GFP localisation which correspond to every major organelle and compartment. We have shown that MT is useful to identify new sequences that determine subcellular localisation as well as known targeting motifs.
Bejarano
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LA
Cell Biology and Biophysics Programme, European Molecular Biology Laboratory, Heidelberg, Germany.
González
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C
D000595:Amino Acid Sequence; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000737:chemistry / Q000235:genetics; D017931:DNA Primers; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics*; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D019151:Peptide Library; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA / Q000379:methods*; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection
D017931:DNA Primers; D008164:Luminescent Proteins; D019151:Peptide Library; D049452:Green Fluorescent Proteins; D004247:DNA