[maker-devel] gff pass thru problem and unsupported EST nucleotides

[Megan]

Maker folks,
I am re-annotating a single contig and I am having a few problems.

First, I am having trouble passing through a Maker derived gff (from Maker 2.09, with some modifications to gene names and functional information added).  The gff file passes the modencode validator but Maker always fails on the first gene in the file, regardless of which gene comes first.  So it appears to be a systematic error across the entire file.  The Maker error is "Check your input GFF3 file for errors! (from GFFDB)".   I have tried Maker 2.10 and 2.31, using both genome_gff with model_pass=1 and pred_gff.  Attached is a gff with the first 2 genes.  

Second, when I updated to Maker 2.31, Maker now complains that my EST fasta file has nucleotides that are not supported [RYKMSWBDHV].  It suggests "set -fix_nucleotides on the command line to fix this automatically".  Is the -fix_nucleotides a Maker flag?  What exactly does it do?  Does it remove the entire sequence or replace ambiguous bases with a randomly selected one?  Half of my 20k ESTs contain these characters, so I don't want to throw them out entirely.  

Also, just curious, has Maker never supported these characters but just never complained?  I used this EST data set with Maker 2.09.  I did note poor EST coverage, but thought it was an issue with the EST data itself.

I appreciate any suggestions.
Thanks,
Megan
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