Augustus/Maker2 failing due to invalid nucleotides in sequence

Question

[harishk0201]

Hi!

I'm trying to annotate genome using Maker. So far, I have the gene predictions using genemark, a repeat masked GFF and augustus trained model.

Now when I'm trying to run Maker2, I have the following error:

/apps/busco/augustus-3.2.3/bin/augustus: ERROR Invalid nucleotide '�' encountered.

/apps/busco/augustus-3.2.3/bin/augustus: ERROR Invalid nucleotide '�' encountered.

ERROR: Augustus failed --> rank=NA, hostname=##### ERROR: Failed while preparing ab-inits ERROR: Chunk failed at level:0, tier_type:2 FAILED CONTIG:Contig8_pilon

ERROR: Chunk failed at level:4, tier_type:0 FAILED CONTIG:Contig8_pilon

I am examining contents of the fasta file and run log. I found this thread, but it honestly doesn't seem to answer what I want.

Any suggestions?

grep -v "^>" contig8.fasta | sed 's/\(.\)/\1\n/g' | sort | uniq -c

2030 A 1123 C 1211 G 1948 T

There are contigs longer than this (some greater than 500kb) and I'm getting the error on all of them, so I think this isn't a length issue.

[Juke34]

Did you try the -fix_nucleotides parameter while running Maker ?

[harishk0201]

Yes I did! I don't know what is the reason behind the error though.

[Juke34]

It looks like the issue is due to an unprintable character. Have you manipulated the file with Windows at some point ?

Answer 1

[Philipp Bayer]

It's hard to fix without looking at Contig8, looks like there's some weird extra character in there.

There are a few threads on Biostars on removing non-nucleotide letters from an input sequence, have a try: C: Remove unwanted characters from FASTA file

Is Contig8 the first contig in the file? That could indicate other problems with the file.

[harishk0201]

No, its the 8th contig. I picked one out of the blue to check if there was something wrong in the contigs.

I've added it to the post, should have done that in the first place.

Thanks for the help!

[Philipp Bayer]

Oh that's strange. Have you tried deleting the maker output folder with the temporary files before rerunning? maybe something's incomplete in there