Validation of predicted anonymous proteins simply using Fisher's exact test.

We introduce the use of the straightforward Fisher's exact test as a postprocessing of the results provided by the popular CODEML sequence comparison software. The respective rates of nucleotide changes at the nonsynonymous versus synonymous position (as determined by CODEML) are turned into entries into a 2 × 2 contingency table, the probability of which is computed under the Null hypothesis that they should not behave differently if the ORFs do not encode actual proteins. Using the genome sequences of two recently isolated giant viruses, we show that strong negative selection pressures do not always provide a solid argument in favor of the existence of proteins.