Methods to study translated pseudogenes: in vitro translation, fusion with a tag/reporter gene, and complementation assay.
The technical challenge in proving that a given expressed pseudogene is in fact translated into a functional protein is specificity. To circumvent this challenge, one approach is to use PCR in order to generate a series of clones that allow expression of the protein of interest either native or fused to a tag, which can facilitate purification, detection, and complementation in both bacterial and mammalian cells. This approach allows an assessment of whether a putative pseudogenic protein possesses enzymatic activity, to identify its subcellular localization and to test its capacity to complement the parental homologue.
Nutritional Genomics Group, School of Biotechnology, Dublin City University, Dublin 9, Ireland, anne.parle-mcdermott@dcu.ie.
Anne
A
Parle-McDermott
D003001:Cloning, Molecular; D015870:Gene Expression; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D023281:Genomics / Q000379:methods*; D006801:Humans; D066298:In Vitro Techniques; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D011544:Pseudogenes / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics
D011993:Recombinant Fusion Proteins