RAPID EXPRESSION SCREENING OF CAENORHABDITIS-ELEGANS HOMEOBOX OPEN READING FRAMES USING A 2-STEP POLYMERASE-CHAIN-REACTION PROMOTER-GFP REPORTER CONSTRUCTION TECHNIQUE

In this paper a description is given of the expression pattern of the Caenorhabditis elegans homeobox gene ceh-38 using GFP reporter constru cts, which were generated using a two-step polymerase chain reaction ( PCR) procedure. This method allows fast analysis of genes of interest by looking at their expression in vivo using their putative promoter r egion to control the expression of a reporter gene. In this case the m ethod was applied to screen C elegans homeobox-containing genes to ide ntify those that are expressed in the head and nervous system. The C e legans genome project has made rapid progress, and more than 79 megaba ses of genomic data with several thousand open reading frames are avai lable. This information can be used to design primers from putative pr omoter regions, which are amplified using long-range PCR. The long-ran ge PCR product is then directly joined to the vector in a long-range F ill-in PCR. Since many genome projects are advancing rapidly, this app roach should also be applicable for other model systems, and the metho d lends itself to automation, since no gel-purification steps are nece ssary. ceh-38 is a member of the ONECUT class of homeobox genes. Expre ssion of ceh-38 starts during embryogenesis. In larvae and adults, exp ression was seen in many different types of tissues, such as the phary nx, gut, hypodermis and many nerve cells. (C) 1998 Elsevier Science B. V. All rights reserved.