In situ hybridization screen in zebrafish for the selection of genes encoding secreted proteins

An in situ hybridization expression screen using a signal sequence trap sys tem has been conducted in zebrafish to isolate cDNAs that encode secreted p roteins. Random clones (secreted expressed sequence tags; sESTs) were seque nced from zebrafish embryonic (18-24 hr postfertilization) and adult kidney libraries. From the two RNA sources, 627 random sEST cDNAs were identified as being homologous or identical to known genes and 166 clones encode curr ently unidentified genes. The sESTs represent a broad range of enzymes and other regulatory molecules. Whole-mount in situ hybridization analysis was carried out by using antisense probes generated from 244 selected sESTs, an d a range of expression patterns was obtained. Genetic mapping undertaken w ith sEST sequences demonstrated that assignment of map position was attaina ble by using 5 ' primers. The signal sequence trap system used in this work has yielded a range of cDNAs that encode secreted proteins and, together w ith analysis of patterns of expression and genetic mapping, has the potenti al to facilitate analysis of signaling pathways central to development and physiology. (C) 2001 Wiley-Liss, Inc.