transmembrane receptor or anchored to the surface(e. g, through a glycosyl phosphatidylinositol phosphate(GPI linkage) Fortunately we usually have the luxury of working with genes that are at least partially characterized by their biological prop- erties. But what about the genes of unknown origin or function? In this new age of genomics, many of the genes we obtain are like" genes, belonging to large families of related genes that share only a minimal percentage of homology with a known gene Despite these similarities there is often no way to know whether the same expression and purification methods used for one ortho- logue or homologue will be effective for another. Thus one is immediately faced with the challenging prospect of having to consider multiple expression strategies in order to get the protein expressed and purified to sufficient levels in an active form, in addition to not knowing what activity to look for Can You obtain the cdnA? Before embarking on an expression project you will need to locate a CDNA copy of the gene of interest. It is also possible in theory to express genomic DNA containing introns, provided that the expression host will recognize the proper splice junctions In practice, however, this is not often the most efficient route to expression because it is not usually known how the introns will affect expression levels or whether the desired splice variant will be expressed. Furthermore most mammalian genes are inter rupted by multiple intron sequences that can span many kilobases in length. This can make subcloning of genomic DNA consider ably more difficult than for the corresponding cDNA The three most common ways to obtain a known gene of interest include purchase from a distributor of clones from the Integrated Molecular Analysis of Genomes and their Expression(image)consortium(http:/image.liNlgov/),requests from a published source such as an academic lab, or RT-PCR cloning from RNa derived from a cell or tissue source. IMAGE clones can be found by performing a BLAST search of electronic database such as Gen Bank, which can be accessed at the National Library of Medicine PubMed browser (http://www.ncbinlm.nih.gov/pubmed/).Fromthereyou quickly determine if a sequence is present, if it is full ler publications related to this gene, and possible sources of the (tissue sources, personal contacts, etc). Most expressed sequence tags(EST's) matching the gene of interest are available as IMAGE clones. The trick is to find one that is full length. It is Eukaryotic Expressiontransmembrane receptor or anchored to the surface (e.g., through a glycosyl phosphatidylinositol phosphate (GPI linkage). Fortunately we usually have the luxury of working with genes that are at least partially characterized by their biological prop￾erties. But what about the genes of unknown origin or function? In this new age of genomics, many of the genes we obtain are “like” genes, belonging to large families of related genes that share only a minimal percentage of homology with a known gene. Despite these similarities there is often no way to know whether the same expression and purification methods used for one ortho￾logue or homologue will be effective for another. Thus one is immediately faced with the challenging prospect of having to consider multiple expression strategies in order to get the protein expressed and purified to sufficient levels in an active form, in addition to not knowing what activity to look for. Can You Obtain the cDNA? Before embarking on an expression project you will need to locate a cDNA copy of the gene of interest. It is also possible in theory to express genomic DNA containing introns, provided that the expression host will recognize the proper splice junctions. In practice, however, this is not often the most efficient route to expression because it is not usually known how the introns will affect expression levels or whether the desired splice variant will be expressed. Furthermore most mammalian genes are inter￾rupted by multiple intron sequences that can span many kilobases in length. This can make subcloning of genomic DNA consider￾ably more difficult than for the corresponding cDNA. The three most common ways to obtain a known gene of interest include purchase from a distributor of clones from the Integrated Molecular Analysis of Genomes and their Expression (IMAGE) consortium (http://image.llnl.gov/), requests from a published source such as an academic lab, or RT-PCR cloning from RNA derived from a cell or tissue source. IMAGE clones can be found by performing a BLAST search of an electronic database such as GenBank, which can be accessed at the National Library of Medicine PubMed browser (http://www.ncbi.nlm.nih.gov/PubMed/). From there you can quickly determine if a sequence is present, if it is full length, publications related to this gene, and possible sources of the gene (tissue sources, personal contacts, etc). Most expressed sequence tags (EST’s) matching the gene of interest are available as IMAGE clones. The trick is to find one that is full length. It is Eukaryotic Expression 497