Using the pcr.seqs() function from the mothur project (https://mothur.org/wiki/pcr.seqs/) I have performed in silico pcr for the full ITS region (ITS1_ITS4) against the Unite database (sh_general_release_dynamic_s_all_10.05.2021). Command below.
pcr.seqs(fasta=sh_general_release_dynamic_s_all_10.05.2021.fasta, oligos=primers_ITS1f_ITS4r_Fung.oligos ,pdiffs=3 ,rdiffs=3)
The results were underwhelming as I only got 55 sequences that matched, out of the 205125 database sequences.
Have some of you experienced this? Is Unite not appropriate for the full ITS region?
Thank you for the help, João Colaço