My 100G metagenome data randomly subsample 10G and 50G data, 10G data can find one ORF, 50G data can't find any ORF. Theoretically, as the sequencing depth increases, more ORF should be found, and 10G species should also be found in the 50G data. However, none ORFs were found in my 50G metagenome data. Why did it happened? Was it because of randomly subsample?