[vmikk]

cutprimers.sh error

Hello Brian!
I tried to extract a region of interest from the SILVA database with cutprimers.sh (from BBMap v.35.66) and encountered the following error:

Exception in thread "main" java.lang.ArrayIndexOutOfBoundsException
	at java.lang.System.arraycopy(Native Method)
	at java.util.Arrays.copyOfRange(Arrays.java:2551)
	at jgi.CutPrimers.process(CutPrimers.java:166)
	at jgi.CutPrimers.main(CutPrimers.java:35)

I split the database and found that only a small part causes this error, but I can not figure out what is wrong with it. I would appreciate your help.

Reproducible example:

msa.sh in=ERR.fasta out=pr1.sam literal="CCGTTCTTAGTTGGTGG"
msa.sh in=ERR.fasta out=pr2.sam literal="TAACAAGGTTTCCGTAGGTG"
cutprimers.sh in=ERR.fasta out=ERR_cut.fasta sam1=pr1.sam sam2=pr2.sam

File is in the attachment.

Edit: Thanks a lot for the fix! With v.35.68 everything works fine!

[Brian Bushnell]

Thanks for finding the specific part that triggers the error, that will be really helpful! I'll try to address that as soon as possible (hopefully tomorrow).

Edit: It's fixed now; I'll upload the new version soon.

[Bioinform]

Hi Brian,

I have a some error with BBsplit.sh (BBmap35.66). I get the following error.,

Module slurm/15.08.1 loaded
java -Djava.library.path=/home/software/BBMap/bbmap/jni/ -ea -Xmx18891m -cp /home/software/BBMap/bbmap/current/ align2.BBSplitter ow=t fastareadlen=500 minhits=1 minratio=0.56 maxindel=20 qtrim=rl untrim=t trimq=6 in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq ref=/home/Whole_genome_analysis/Reference/NCPPB382/Cmm382_genome.fasta,/home/Whole_genome_analysis/Reference/NCPPB382/Xcitri_jx6.fasta basename=PST_407_%.fq outu=PST_407_unmapped.fq
Executing align2.BBSplitter [ow=t, fastareadlen=500, minhits=1, minratio=0.56, maxindel=20, qtrim=rl, untrim=t, trimq=6, in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq, ref=/home/Whole_genome_analysis/Reference/NCPPB382/Cmm382_genome.fasta,/home/Whole_genome_analysis/Reference/NCPPB382/Xcitri_jx6.fasta, basename=PST_407_%.fq, outu=PST_407_unmapped.fq]

Converted arguments to [ow=t, fastareadlen=500, minhits=1, minratio=0.56, maxindel=20, qtrim=rl, untrim=t, trimq=6, in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq, basename=PST_407_%.fq, outu=PST_407_unmapped.fq, ref_Cmm382_genome=/home/Whole_genome_analysis/Reference/NCPPB382/Cmm382_genome.fasta, ref_Xcitri_jx6=/home/Whole_genome_analysis/Reference/NCPPB382/Xcitri_jx6.fasta]
Creating merged reference file ref/genome/1/merged_ref_1588668246460521.fa.gz
Ref merge time: 1.421 seconds.
Executing align2.BBMap [ow=t, fastareadlen=500, minhits=1, minratio=0.56, maxindel=20, qtrim=rl, untrim=t, trimq=6, in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq, outu=PST_407_unmapped.fq, ref=ref/genome/1/merged_ref_1588668246460521.fa.gz, out_Cmm382_genome=PST_407_Cmm382_genome.fq, out_Xcitri_jx6=PST_407_Xcitri_jx6.fq]

BBMap version 35.66
Set MINIMUM_ALIGNMENT_SCORE_RATIO to 0.560
Retaining first best site only for ambiguous mappings.
NOTE: Deleting contents of ref/genome/1 because reference is specified and overwrite=true
Writing reference.
Executing dna.FastaToChromArrays2 [ref/genome/1/merged_ref_1588668246460521.fa.gz, 1, writeinthread=false, genscaffoldinfo=true, retain, waitforwriting=false, gz=true, maxlen=536670912, writechroms=true, minscaf=1, midpad=300, startpad=8000, stoppad=8000, nodisk=false]

Set genScaffoldInfo=true
Writing chunk 1
Set genome to 1

Loaded Reference: 0.005 seconds.
Loading index for chunk 1-1, build 1
No index available; generating from reference genome: /home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/ref/index/1/chr1_index_k13_c7_b1.block
Indexing threads started for block 0-1
Indexing threads finished for block 0-1
Generated Index: 4.556 seconds.
Analyzed Index: 2.561 seconds.
Started output stream: 0.526 seconds.
Cleared Memory: 0.209 seconds.
Processing reads in single-ended mode.
Started read stream.
Started 16 mapping threads.
java.lang.ArrayIndexOutOfBoundsException: 0
at stream.FASTQ.quadToRead(FASTQ.java:723)
at stream.FASTQ.toReadList(FASTQ.java:653)
at stream.FastqReadInputStream.fillBuffer(FastqReadInputStream.java:111)
at stream.FastqReadInputStream.nextList(FastqReadInputStream.java:96)
at stream.ConcurrentGenericReadInputStream$ReadThread.readLists(ConcurrentGenericReadInputStream.java:659)
at stream.ConcurrentGenericReadInputStream$ReadThread.run(ConcurrentGenericReadInputStream.java:638)
Exception in thread "Thread-22" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-23" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-17" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-21" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-25" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-30" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-16" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Detecting finished threads: 0, 1Exception in thread "Thread-19" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-20" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-28" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-29" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-26" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-18" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
, 2, 3, 4, 5, 6, 7Exception in thread "Thread-27" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
Exception in thread "Thread-24" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
, 8, 9, 10, 11, 12, 13, 14Exception in thread "Thread-31" java.lang.NullPointerException
at align2.AbstractMapThread.run(AbstractMapThread.java:563)
, 15

**************************************************************************
Warning! 16 mapping threads did not terminate normally.
Check the error log; the output may be corrupt or incomplete.
Please submit the full stderr output as a bug report, not just this message.
**************************************************************************




------------------ Results ------------------

Genome: 1
Key Length: 13
Max Indel: 20
Minimum Score Ratio: 0.56
Mapping Mode: normal
Reads Used: 3200 (446131 bases)

Mapping: 2.454 seconds.
Reads/sec: 1303.92
kBases/sec: 181.79


Read 1 data: pct reads num reads pct bases num bases

mapped: 0.9688% 31 0.7738% 3452
unambiguous: 0.3438% 11 0.3373% 1505
ambiguous: 0.6250% 20 0.4364% 1947
low-Q discards: 0.0000% 0 0.0000% 0

perfect best site: 0.0313% 1 0.0043% 19
semiperfect site: 0.0313% 1 0.0043% 19

Match Rate: NA NA 85.1230% 2941
Error Rate: 17.0455% 30 14.8770% 514
Sub Rate: 17.0455% 30 14.4718% 500
Del Rate: 1.7045% 3 0.0868% 3
Ins Rate: 3.4091% 6 0.3184% 11
N Rate: 0.0000% 0 0.0000% 0
Exception in thread "main" java.lang.RuntimeException: BBMap terminated in an error state; the output may be corrupt.
at align2.BBMap.testSpeed(BBMap.java:489)
at align2.BBMap.main(BBMap.java:34)
at align2.BBSplitter.main(BBSplitter.java:46)

Is there any thing you help with?

Thanks

[GenoMax]

I think you are running out of memory (@Brian will confirm). How much RAM do you have (looks like you are asking for ~18G)?

[Bioinform]

Thanks for the quick response. I am running it in farm cluster with --mem 20000. I think it should not be the memory thing.

[GenoMax]

Job is starting 16 threads yet I don't see you asking for threads=16 in your command line. Is that right?

[Brian Bushnell]

BBMap autodetects the number of processors available and uses all of them, by default. This looks to me like a corrupt fastq file (GGAGC_trimmed1.fastq). I'll have to look at the specifics... but probably, either some reads were trimmed to zero length and I am not correctly handling that (so it is a bug), or the file is corrupt and some other lines (like the header or the line with the +) are zero length.

I'll get back to you some time later today. If it's a bug due to zero-length reads, I'll post a fixed version on Monday, but in the mean time you can circumvent that by discarding zero-length reads when trimming, or trimming to a minimum length of 1 (which is what BBDuk does, to avoid zero-length reads).

[Bioinform]

I used as batch with following

#!/bin/bash -l
#SBATCH -J bbsplit
#SBATCH -o myjob.%A_%a.output
#SBATCH -e myjob.%A_%a.error
#SBATCH -p serial
#SBATCH -t 999:00:00
#SBATCH -N 1
#SBATCH -n 8
#SBATCH --mem 20000

echo start

bbsplit.sh in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq ref=/home/spthapa/Whole_genome_analysis/Reference/NCPPB382/Cmm382_genome.fasta,/home/Whole_genome_analysis/Reference/NCPPB382/Xcitri_jx6.fasta basename=PST_407_%.fq outu=PST_407_unmapped.fq

echo finished

I also ran directly using
bbsplit.sh in=/home/Whole_genome_analysis/HiseQ/HiseQ_data/PST/Trimmed_Seq/GGAGC_trimmed1.fastq ref=/home/spthapa/Whole_genome_analysis/Reference/NCPPB382/Cmm382_genome.fasta,/home/Whole_genome_analysis/Reference/NCPPB382/Xcitri_jx6.fasta basename=PST_407_%.fq outu=PST_407_unmapped.fq

I get the same error.

I am not sure of that.

Thanks

[GenoMax]

@Brian: @Bioinform appears to be submitting under slurm so I am surprised that it is allowing BBMap to grab all cores (perhaps @Bioinform is specifying them on other part of the command line).

The error appears to be happening in first few reads if I am reading the stats right.

EDIT: @Bioinform: Add threads=8 to match your request of 8 cores (-n 8) via slurm (that won't fix the problem but just to be consistent).

[Bioinform]

@ GenoMax

Thanks, will match it.

@Brian

with bbduk.sh also i am getting error messages.

Module slurm/15.08.1 loaded
java -Djava.library.path=/home/software/BBMap/bbmap/jni/ -ea -Xmx6519m -Xms6519m -cp /home/software/BBMap/bbmap/current/ jgi.BBDuk2 in=GGAGC_trimmed2.fastq trimq=6 out=407_PST.fastq
Executing jgi.BBDuk2 [in=GGAGC_trimmed2.fastq, trimq=6, out=407_PST.fastq]

BBDuk2 version 35.66

NOTE: No reference files specified, no trimming mode, no min avg quality, no histograms - read sequences will not be changed.
Initial:
Memory: max=6550m, free=6482m, used=68m

Input is being processed as unpaired
Started output streams: 0.054 seconds.
java.lang.ArrayIndexOutOfBoundsException: 0
at stream.FASTQ.quadToRead(FASTQ.java:723)
at stream.FASTQ.toReadList(FASTQ.java:653)
at stream.FastqReadInputStream.fillBuffer(FastqReadInputStream.java:111)
at stream.FastqReadInputStream.nextList(FastqReadInputStream.java:96)
at stream.ConcurrentGenericReadInputStream$ReadThread.readLists(ConcurrentGenericReadInputStream.java:659)
at stream.ConcurrentGenericReadInputStream$ReadThread.run(ConcurrentGenericReadInputStream.java:638)
Exception in thread "Thread-16" Exception in thread "Thread-15" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-19" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-4" Exception in thread "Thread-11" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-8" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-5" Exception in thread "Thread-17" Exception in thread "Thread-7" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-6" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-12" Exception in thread "Thread-9" Exception in thread "Thread-10" Exception in thread "Thread-13" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Exception in thread "Thread-18" java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
java.lang.NullPointerException
at jgi.BBDuk2$ProcessThread.run(BBDuk2.java:2417)
Processing time: 0.110 seconds.

Input: 5200 reads 724560 bases.
Result: 5175 reads (99.52%) 724439 bases (99.98%)

Time: 0.176 seconds.
Reads Processed: 5200 29.58k reads/sec
Bases Processed: 724k 4.12m bases/sec
Exception in thread "main" java.lang.RuntimeException: jgi.BBDuk2 terminated in an error state; the output may be corrupt.
at jgi.BBDuk2.process(BBDuk2.java:888)
at jgi.BBDuk2.main(BBDuk2.java:70)

Thanks

[Brian Bushnell]

OK, I looked at the code - your fastq file is corrupt; you are missing a "+" symbol on a line. It's probably truncated. You should go back to the raw reads and reprocess them.

@Brian: @Bioinform appears to be submitting under slurm so I am surprised that it is allowing BBMap to grab all cores (perhaps @Bioinform is specifying them on other part of the command line).

Oh, that's good to know; I've never used Slurm.

[Bioinform]

Thanks Brian. Have a gr8 weekend.

[Bioinform]

Are you talking about some lines, my fastq file do have + symbol.

This is my fastq file

@DJB77P1:527:H7RDTADXX:1:1101:2830:2226 1:N:0:
AATTTCCGTCACCCTTTTAAGTCCCCCAGGCGGGGGGCTCGACGAGAGCGACAGACCTTGTGTGTAGAAGTTTCAAAATGCTTATGCATCAAGAGACAGTGCCCTGCCCGAAGATATTTACATTCGGTGTGCCTTGGGCGTATA
+
FFHHHHHJJJJJJJJJJJJICHIIJJIJJIJJJJDDBDDDDBDDBBDDDDDDDDDDDDDDCC??BCCDDDACDDEDDDDDDDEDDDDEECDDDDDDCDDCCDDDCBDBDD<@B<ACDEEDEDDEDED?BBDDDC@>CBBDBB@C
@DJB77P1:527:H7RDTADXX:1:1101:2830:2226 3:N:0:
AGCTCGCCTTACGATGCTCGCACACGAAGACCGCAGAATTAACCAATAACTCCCTGCCTCAAGCCTTGGGCGTATACGCCCAAGGCACACCGAATGTAAATATCTTCGGGCAGGGCACTGTCTCTTGATGCATAAGCATTTTGAAACT
+
=DFFFFHHHHHJJJJJIJJJJJJJJJJJGIJJJJJJJJHHHHHHFFEFFDDEEDEDDDDDDDDDDDDCCDDDDDDDEDDDDDDDDDDDDDDDDDDDCDEEEEDDEEEBDDDDDDDDDDDDDDDDDDDDCDDDDDDDCDDDDECDDDDD
@DJB77P1:527:H7RDTADXX:1:1101:4220:2204 1:N:0:
CGACGCCTGCATAAGGGCTTTCGCTGATTGCCGCGCCCAGGCCCAGGCTGAAATCGCCCGGTTCAGCGTGGGCGAACGGCAACGACAGGGCAGGCAAAGCAACAACGGTCAGTGCGGGAAGGAAAAAGTGTTTCATGACGGCGG
+
DDHHGHGJIJJJJJHIJJJJJJIIIJJJEIIIGIEIJHHEFDFDDDDDDDDDDDDDDDD@B5;<ACC9@BD@?BD<BDDDBDBDBBDBDB<?BBDDBDCDDDBDDDDD@@BD@ACDDDDDDBDDDDDD:::@CC@C@CDBBBBD
@DJB77P1:527:H7RDTADXX:1:1101:4220:2204 3:N:0:
AGCTTATACACGCCAGAAAGATCACTCAGAGAGCCGCCGTCATGAAACACTTTTTCCTTCCCGCAC
+
=DDDFFHHHGGIIIIJJJJJIJIIIIIJFIIIJJJIJIJIFGIJIJJGGIHGHGHGFFFEFFDDDD
@DJB77P1:527:H7RDTADXX:1:1101:4290:2210 1:N:0:
CTGTACTCGATCGGCAAGGATTCGGCCGTGATGCTGCACCTGGCGCGCAAGGCTTTCTTCCCCGGCAAACTGCCATTCCCTGTGATGCATGTCGATACCCGCTGGAAATTCCAGGAGATGTATCGCTTTCGCGACCAGATGGTC
+
FFHFDHHJIBGIGIGIJJJIJJIIJIGIDGBHIIIGEAHIGJGGAHFDDD?BDDDC>CACCDCDDDBBDDDDDDDCCADDDDCDEECDDEC@CAB?ABDABD<BBDCCDCDDDCDBDBCCC>@BBB@D0><9<>BDDD8@ACCC
@DJB77P1:527:H7RDTADXX:1:1101:4290:2210 3:N:0:
TCGACATGCATCACAGGGAATGGCAGTTTGCCGGGGAAGAAAGCCTTGCGCGCCAGGTGCAGCATCACGGCCGAATCCT
+
?BDA@BBCCDEDDDDDDDDDDDDDDDDDEDDDDDDDBBDDDDDDDDDDDDDDDDDDD:A>CCABDDDDDDDDDDDDDCC
@DJB77P1:527:H7RDTADXX:1:1101:6926:2244 1:N:0:
ATATCGTATTTCCAGAGTGCCACTGGCTGATAAAGAAAGAGGCCGGGCATATGCTGATTCTGGCTATTTACCTTGACGCTCGCAGCAGCGATATCGACGGATCAGGGTTTAAATCATCCTCGTATTTAACTTTGGGAAAACAAC
+
FFGHHHHJJJJJJJJJJFHIJJJJJIJJJIJJJIIIJJGIIJIJIJJJIIJJJJIHHHHGHHFFFFFEEEEEEEDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDEDDDDDDDD<BDDEDEDDEDDCDDDDDDDDD
@DJB77P1:527:H7RDTADXX:1:1101:6926:2244 3:N:0:
GGAGCTACCTTCCTGAACCAGGCGCATCACGTGTGTTTTTGACACCGATAGTTGTTTTCCCAAAGTTAAATACGAGGATGATTTAAACCCTGATCCGTCGATATCGCTGCTGCGAGCGTCAAGGTAAATAGCCAGAATCAGCATATGCCC
+
CCCFFFFFGHHHHJJJJJJIGIJJJJIIJJJHHGGHIJJJJJJJIIJGIIFHHHHFFFFFDEEEDEDDEDDECDDDDDDDDEEEDEDDDDDDDDDDDBDDDDBDDDDDDDDDDDDDDDBDDDDD>>CC@DACDDDDDDDDDDDCCDECDD
@DJB77P1:527:H7RDTADXX:1:1101:9747:2206 1:N:0:
TGCTCGTCAGGTACTCGCTGAGGTTGTACAACTGCGCGATGTCGTTGCCATCGGCCATCAACACGCTACTGCGTACGTAGCCGCCGTTGAGCCACTCAAAGCTGTCGTTACCGAAGCTTGCGCGTATTTCTCCACGAACTTCGC
+
FFHHHHHJJIJIHIJJJJJJJJJJJJJJIJJJJJGJJJJJJIJIJHEHHHFFFDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDBDDDDDDBDDDBDDDEEEDDDDDDDDDDD?B
@DJB77P1:527:H7RDTADXX:1:1101:9747:2206 3:N:0:
AACGTCTTCGAGCTTTCTGGCGGCAGCATTATCAACAATCTGGTCGCCGGGTTTGGCCGGGACACGATTACAGTGTCGG
+
8@<@<ACBDDDDBDDDDDCDCDDBDDDDCDCCCDDCCDC?ACC@CBDDDDDBBD?B>CDDD>BDBD3<?<@AC>>@@BB
@DJB77P1:527:H7RDTADXX:1:1101:12008:2199 1:N:0:
CACTGCGGCCGCCTGCGATGAAGTTCTGGCCGAGAATGTTAGGCATCGGTATCTCCTGTTGTTTGATGTTGGAGGCGGGCGCCGAAGGGATCCGGACCGTGCGCTCCTTATGGGTTGCCGAGTTGCTTTGCGGGGTGGCACTGG
+
FFHHHHHJJJJJJJJJJHJJJJJJJJJJJJJJHJIHHHHHHFFFFFEDDDDDDFDDDDDDDDDDDCDDDDDDCDDDDCDBBDDB9B9BDB<<C<B>@@.5?B>@B>@CC>C>CD3<9<A9<B@BCA>ACAC<>BD9BD@A@<C@
@DJB77P1:527:H7RDTADXX:1:1101:12008:2199 3:N:0:
AGCTTAAACAGCAGTGGCGACATAAAGGCTAAACGCTGTCTGGCAGTTGCGTCAGCAGCGAATCAA
+
=DDFFFHHHHHJIIIJJJJJJJJJJJJJGIIJJJJIIJGIGIIIEIIIIJJJHFHFFFFDDDDDDD
@DJB77P1:527:H7RDTADXX:1:1101:14057:2199 1:N:0:
AGCCGGAAATAAGCAACGTCCGATCAATCCTGTCACAGGGTTTGGTGGCAGTGATCGCGCTGGCTCTCGATGGTGAAGGTTCAATGAGGCATTTGAAGATCAAGAGCCTTTCCCTTACCGGACATGACGGACCCGGTATACGCG
+
FFHHHHHJIJJJJJJJJFHIIJIJJJJJJJJJJJJJJJJJHIJJJIHHHHFFFFFEDDDDDDDDDDDDDDDEDACDDDDCCDDEEDDDDDDDDDEEDDDDDDDDDDDDDDDDDDDDDDDDDDBDDDDDDDDDDDDDBDDEEDDD
@DJB77P1:527:H7RDTADXX:1:1101:14057:2199 3:N:0:
AGCTATTGATATTCCAAATCTTGGGTAAGCATCACGATCTGATTGGATCGGATCTTCTCGCGTATA
+
=DDFFFHHHHHJJJJJJJJJJJJJJHHIJJJJJ@GIJIJJJIJJJJIJHIJJJJIJJJIIJJHHFF
@DJB77P1:527:H7RDTADXX:1:1101:14176:2213 1:N:0:
CCTTGGCTTCGGCGTGCGCCGAGAAGGCGGGGATCATCAAGGCCAGAAAAATGGCATGAAATTTCATCGGTGCTCCTGCGGATGAATATGACTTCATTGGATCCAAAATTTACTCCAGTCGCGTGCCTCTGACTAGCGGGTGCC
+
FFHHHHHJJJJJJJIJJJJJJJJJJJJJJHDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDEEEDDBBDDDDDDDDDBDDDDEEEEEDDDDDEEDDDDDDDDDDDDDDEDDDDDDDDDDDDDDDDDDDDDCCDDDDBBBD
@DJB77P1:527:H7RDTADXX:1:1101:14176:2213 3:N:0:
AGCTAGAACAACAGCGATTGCCGGAATCAACCCGCGCCAGGCAAGGTCAGCCAACATGACAACCCGACGAAAGGCCACTGGCACCCGCTAGTCAGAGGCACGCGACTGGAGTAAATTTTGGATCCAATGAAGTCATATTCATCCGCAG
+
=DDFFFHHHHHJJIJIJJJJJJJJJJJJJJJJJJJJJJJJHHHHEF@DFEEEEEDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDCDEDDDDDBDDDDDDDDDDD>CCDDEDDDDDDDDDDDDDDDCDEEEEEEEEEDDDDC
@DJB77P1:527:H7RDTADXX:1:1101:15053:2181 1:N:0:
CAGCTCAGGAGGATTTTCCCGCGTGTCGGCCACAGCAAAGGACACGCCCCGGTTCGCCAGGAAGCGAACCAGGGACATGCCACTCTTGCCAAGCCCCACAACAATGCGGAAGCGGTCGGAAACGATCAGGGACACTCGTTCTAC
+
FFHHHHHJJJJJJJJJJJJJIJJJJJJJJJJJJJJJJJJIJJJHHHFFDDBD?BDDDDDDDDDDDDDDDDDBDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDCDDDDDDDDBDDDBDDDDDDDDDDDDDDDCDDDDDDED
@DJB77P1:527:H7RDTADXX:1:1101:15053:2181 3:N:0:
CAGTGGTCATTCAGGTTGCGTCTTTCAA
+
<@@?@?@?@???????????????????
@DJB77P1:527:H7RDTADXX:1:1101:15413:2202 1:N:0:
GTGCAATTGCCGACCTGCTGGTGGTGGACGGCGACCCGCTGAGCGATATCAGCTGCCTGGTGGGCCAGGGCGAGCAGTTGGCGATGATTGTTCAGGGTGGTCATGTGCACAAGAACACCCTGGCCTGATCAGCCGTCAGTCGGC
+
FDHGHHHIIIJJIJJIJJIJGFHIDHIHIIJJIJJHFEDDD@DDDDDDDDEEDDDDCDDD@CDDDDDDDDBDDDDDDCCDDDBBDDDEEDDDDEDDDCDD>CCDDCDDEDDDDDDDDDDDDDDDDDDDDDEDDDDBBBDDDBBB
@DJB77P1:527:H7RDTADXX:1:1101:15413:2202 3:N:0:
CAGGCCAGGGTGTTCTTGTGCACATGACCACCCTGAACAATCATCGCCAACTGCTCGCCCTGGCCCACCAGGCAGCTGA
+
?BDDDDDDDD<?@DDEDDDDDEDDDDDDDDBDDDCDDDCCDCDDDDBDDDBDCDDDBBBDBBDDDDDDDDBBDDDDDCC
@DJB77P1:527:H7RDTADXX:1:1101:15955:2211 1:N:0:
GTGGCTTCCGTTCCGCCTCAAGGTGGTCGCAAGCATCCGCAGCAGGAATTCCTGCAGGTGGACACGCGCAACATCCTCTTCATATGCGGTGGCGCCTTCTCGGGCTTGGAAAAGGTCATTCAGAACCGTTCCACCCGTGGCGGC
+
FFHHGHHIGIIIJHIIJIJJJIICGGHIIIJIIIIJJJJJJIGIFHGHFFEFFEEEEEEDDDDDDDDDDDDDBDCDDDDDDEEEEEDDDBDBDDB>BDDDDDDDDDDDDDDCCDD@CDDEEECDDDDDBBDDDDDDD<@BDBDD
@DJB77P1:527:H7RDTADXX:1:1101:15955:2211 3:N:0:
AGCTCTCCTTGCTGCGAACTTCGGCGTTGAAACCGATGCCGCCACGGGTGGAACGGTTCTGAATGACCTTTTCCAAGCCCGAGAAGGCGCCACCGCATATGAAGAGGATGTTGCGCGTGTCCACCTGCAGGAATTCCTGCTGCGGATG
+
=DDFFEHFHHHJJJJJGHIIJIIIJJJJJIIJIJJIGIJJJHFFFDDD9?B?CCBD?BBCDDDDDDDDDDDDDCEDDDDDDDDDDDDDDDDDDDDDDDDACDEDABDBDDDDDCBDDDDDDDDDDDDDDDDBDDDDDDDDDDDDDDDD
@DJB77P1:527:H7RDTADXX:1:1101:16246:2219 1:N:0:
CTATATAGACCTGCGCTCTTGGCCAAGTCGATGCCTTGCCGCTGTCGCTCTCGCCGGTCCTCGAAGTCGTCACGCGCTATTTGCAGTGCCACACGCAACAGCATGTCTTGTACGCCTTGCAACACCACCTTGGCGACGCCACTG
+
FFHHHHHJJJJJJJJJJJJJIIIJJJJJJJJJJJJJJJJJJJJJJJJJJJJHHHFDDDDDDDDDDDDDDDDDDDDDDDDDDEDDEDDDDDDDDDDDDDDDDDDDDDDEDDDDEDDDDDDDDDDDDDDDDDDCDCDDDDDDDDCC
@DJB77P1:527:H7RDTADXX:1:1101:16246:2219 3:N:0:
GGAGCTGTGAAGTCGTCATCGCTGAGAAGATCGACCGAATCAGCCGTCTTCCCTTGGTCGAAGCAGAAAGGCTTGTGGACGCGATCAAGGCTAAGGGCGCACGCTTGGCGGTGCCAGGCATCGTCGATTTATCGGAATTGGCCGAGGCAT
+
CC@FFFFFHHHHHIJJJJJIJJJJJJJJJJJJJJJJJJJJIJJJJJJJJHGHHHFFFFFDDDDDDDDDDDDDDDDBDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDDBDDDBDDDDDDDDDCDBDD>BDDCDCBD?B?CC@CDD<BBDBC
@DJB77P1:527:H7RDTADXX:1:1101:18918:2216 1:N:0:
GGGCCGTCCGGGGCAGGAAAAGAAGTCATCGCCCAGCGCTTGCATCGGCTTGGGGTGAACCCGCTCCATCCCTTTGTGGATATCAACTGCGCTGCATTGCCAGCACACCTCATTGAAGCAGAGCTATTCGGTCATAGCCGAGGT
+
FFHHHHDHHHIJJJJJJIIJJJJJJGIJJJJJJJHHHFFCDDCCDDDDDDDDDDD)5?C@BD5955;::>AB:ACC@?B:CDDCDCC:>C<><<>>CDACC>@?9???BCA:>@:4@>AC9?8AACCC>9A8<5>B@CC959@9

[GenoMax]

That offending sequence may be somewhere around #32. Look further down in the file. Otherwise use BBDuk to retrim your data as suggested by Brian to ensure there are no zero length reads.

[babine]

Guidance for mapping with BBMap?

Dear Brian, dear users,

I have reads from an exon based sequence capture and I'm trying to map them to not-so-accurate reference sequences. I started to use BBMap because I hoped it would have a feature that I'm looking for.

My problem is that I'm getting misaligned reads, especially in the end of the reads, when two "stacks" of reads are coming in contact although they should not be but my reference sequence artificially bring them close together. While I would gladly inspect the alignments for those regions and introduce gaps in the reference, I can not do it for more than a few loci. So I was wondering if there is not a more clever way by tweaking the parameters.

I tried a bit with minid, k, padding... but without much success.

Do you have any suggestions on what parameter would have the most impact? Any particular strategy?
Or let me know if you think another algorithm would do it...

Any comment is appreciated...

Cheers