A neo was inserted in reverse orientation to disrupt the reading frame within the coding region. A stop codon was engineered into the proximal neo linker to prematurely truncate the transcript subsequent to Gly-190 in the second extracellular loop. RT-PCR verified presence of the recombinant transcript in mutant brains. Immunohistochemistry using peptides from the region downstream of the stop codon showed that mutants did not produce the corresponding protein. (J:95908)