Enhancer trap screens would be a powerful way to study gene function, as they could simultaneously report gene expression patterns, loss-of-function mutant phenotypes and sequence of the encoded gene products. We are currently developing an approach to perform enhancer trapping, using
cul-1 as a marker to select for random integration of extrachromosomal arrays.
cul-1 is a cell cycle repressor, and cul- 1(null) cells undergo excessive rounds of cell division.
cul-1(null) animals expressing
cul-1(+) from an extrachromosomal array have low brood sizes, because loss of the extrachromosomal array is common and generates a mosaic clone that has excessive cell growth and reduces brood size. When the
cul-1(+) extrachromosomal array integrates into the chromosome, the
cul-1(null) transgenic animals have much higher brood sizes because the array is attached to a chromosome and cannot be lost spontaneously. A large number of random integrants can be obtained by irradiating a transgenic line containing a
cul-1 extrachromosomal array, and then selecting for integrants that rapidly outgrow other worms on a plate. We have used this approach to integrate a lacZ enhancer trapping vector (designed by Andy Fire) into random chromosomal sites. So far, we have generated 155 independant integrants, and 65 (42%) of these integrants express lacZ. Generally, the integrants express lacZ in a pattern that is similar among individuals from the same integrated line, but different between integrated lines. We used a PCR based assay to show that 3 of the integrated lines may be recessive lethal, and we are mapping and characterizing their lethal phenotypes. We are in the process of cloning the endogenous locus for these 3 enhancer traps with lethal phenotypes. Future experiments will show whether the lacZ expression patterns and mutant phenotypes for these three integrants match those of the endogenous genes that contain the enhancer trap insertion.