[maker-devel] Unable to reproduce MAKER blastn results

[Lior Glick]

Hello,

I am running MAKER 2.31.10 with a very simple configuration, with only EST
evidence and the est2genome option enabled (basically a lift-over
procedure).
I noticed that some of my transcripts are not included in the annotation
output and when I looked at the blastn results the reason was clear - they
do not pass the coverage cutoff defined in maker_bopts.ctl. Interestingly,
when I tried running blastn myself, using the same command (taken from the
maker log) and the same blastn version, I got slightly different results.
Specifically, for some of the transcripts the MAKER blastn run produced
less HSPs than my blastn run, resulting in a lower total coverage. The
additional HSPs seem to have good % identity and E-values, so I don't
understand why and how they are discarded. Are the blastn results changed
by MAKER in subsequent steps (after the blastn run)?
Please find attached blastn results from MAKER run and from my run. You can
look at transcript AT1G01740.3 as an example. in my.blastn, there are 8
HSPs, while MAKER.blastn only has 3 of them.
Can you explain the difference? Maybe it has to do with repeat masking or
other processing of the genome sequence?

Just to make sure you have all the details:
Relevant maker_bopts parameters:
pcov_blastn=0.7 #Blastn Percent Coverage Threhold EST-Genome Alignments
pid_blastn=0.85 #Blastn Percent Identity Threshold EST-Genome Aligments
eval_blastn=1e-10 #Blastn eval cutoff
bit_blastn=40 #Blastn bit cutoff
depth_blastn=0 #Blastn depth cutoff (0 to disable cutoff)

Blastn command:
blastn -db
/groups/itay_mayrose/nosnap/liorglic/Projects/PGCM/output/A_thaliana_pan_genome/PGC_de_novo/RESULT_RG_new/per_sample/col-0/liftover_SRR1945757/chunks/chunk00.fa/TMP/maker_sPf3Rf/TAIR10_longest_trans%2Efasta.mpi.10.0
-query
/groups/itay_mayrose/nosnap/liorglic/Projects/PGCM/output/A_thaliana_pan_genome/PGC_de_novo/RESULT_RG_new/per_sample/col-0/liftover_SRR1945757/chunks/chunk00.fa/TMP/maker_sPf3Rf/0/chunk00.0
-num_alignments 10000 -num_descriptions 10000 -evalue 1e-10 -word_size 28
-reward 1 -penalty -5 -gapopen 5 -gapextend 5 -dbsize 1000 -searchsp
500000000 -num_threads 10 -lcase_masking -dust yes -soft_masking true
-show_gis -out

Thank you!
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