Summary of failed rescue attempts from RT-PCR-based clone recovery. Of 107 genes nonrescued to date, 38 were declared failures due to the lack of expected-size PCR amplicons. The cloning process failed for two genes. For 67 genes, clones representing expected-size amplicons were generated. Of these, clone insert sequences of 40 matched a RefSeq sequence other than the targeted gene; 32 of these contained sequences for the correct PCR primers used, whereas the remaining eight did not. Of 27 genes where the clone sequences matched the targeted gene, two failed due to various nonvalidated errors. Eight failed due to technical errors, such as primers amplifying within the ORF. For 17 genes, however, at least half of the clones could not be rescued due to a common unvalidated change. We suggest that clones in the last category may not be true failures, but rather novel splice variants or real polymorphisms that should be considered biologically valid.
