Error in seq.default(from = 1, to = CNt, by = 1) : 'to' cannot be NA, NaN or infinite

Amit Goyal

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Mar 29, 2017, 9:35:00 PM3/29/17

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Hi,

I am trying to analyze the CNV using Tumor:Normal pair using sequenza as recomended in EBI-Cancer-workshop-CNV (https://github.com/mbourgey/EBI_cancer_workshop_CNV)

Below is my commands:

R

library("sequenza")

data.file = "out.small.seqz.gz"

# out.small.seqz.gz is generated by sequenza-utils.py

seqzdata = sequenza.extract(data.file)

CP.example = sequenza.fit(seqzdata)

sequenza.results(sequenza.extract = seqzdata , cp.table = CP.example, sample.id = "C0053", out.dir="/path/to/outdir/sequenza")

# The above command is throwing error

Error in seq.default(from = 1, to = CNt, by = 1) :

'to' cannot be NA, NaN or infinite

In addition: Warning messages:

1: In (function (..., deparse.level = 1) :

number of columns of result is not a multiple of vector length (arg 3)

2: In max(segments$CNt, na.rm = TRUE) :

no non-missing arguments to max; returning -Inf

Is tehre any recomemndation to solve this error?

Thanks,

Amit

Francesco Favero

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Mar 30, 2017, 5:15:56 AM3/30/17

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Hi Amit,

Thanks for the link, it’s a great tutorial!

From the error I see that something is wrong in the object you are passing to sequenza.results, but there are too few details to really help you with.

could you save the R objects (seqzdata and CP.example, or the full R env, if it’s not too big) and send it in attachment.

Thanks!

Francesco

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Amit Goyal

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Mar 30, 2017, 10:10:25 PM3/30/17

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Hi Francesco,

I tried to save the seqzdata and CP.example objects. I am not sure I saved in correct way since I an still new to R.

Also, I am attaching the initial seqz.gz files which I imported as data.file object.

Here, out.small.seqz.gzwas generated using the Normal:Tumor pair bam files for 50kb bin using sequenza-utils.py script.

Thanks,

Amit

Francesco

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CS15_sequenza_cp_table.RData

CS15_sequenza_extract.RData

out.small.seqz.gz

Francesco Favero

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Mar 31, 2017, 4:51:27 AM3/31/17

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Hi Amit,

I had a run with your files, and the problem is that there are very very few heterozygous position in the seqz file.
How did you run the sequenza-utils programs?

Thanks

Francesco

> On 31 Mar 2017, at 04.10, Amit Goyal <amit...@gmail.com> wrote:
>
> <out.small.seqz.gz>

Amit Goyal

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Mar 31, 2017, 4:58:30 AM3/31/17

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I generated the seqz file using the commands given in https://cran.r-project.org/web/packages/sequenza/vignettes/sequenza.pdf file.

Pileup:

samtools mpileup −f hg19.fasta −Q 20 normal.bam | gzip > normal.pileup.gz

samtools mpileup −f hg19.fasta −Q 20 tumor.bam | gzip > tumor.pileup.gz

Generating a genome-wide GC content file:

sequenza−utils.py GC−windows −w 50 hg19.fa | gzip > hg19.gc50Base.txt.gz

Generate a seqz file

sequenza−utils.py pileup2seqz −gc hg19.gc50Base.txt.gz −n normal.pileup.gz −t tumor.pileup.gz | gzip > out.seqz.gz

And last biining:

sequenza−utils.py seqz−binning −w 50 −s out.seqz.gz | gzip > out small.seqz.gz

Amit

Amit Goyal

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Mar 31, 2017, 5:01:14 AM3/31/17

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I think, I didn't provided the target_capture.bed file used to generate the sequencing data during samtools mpileup step as well as to the sequenza−utils.py script.

Do you think it might br the reason?

Amit

Francesco Favero

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Mar 31, 2017, 6:08:45 AM3/31/17

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this really depends on the BAM files used. If you didn’t provided the bed file, it should only take more data.

This shouldn’t impact the number of heterozygous positions.

Is the sequencing from a panel of targeted sequencing?

If this is the case you could try to play around with the mufreq model, even if the method is not very well tested and documented.

The error is caused by the lack of BAF position. I have solved the issue in a new version I’m working on, but it’s not ready to be released, and I don’t recommend you to try it out (since many other problems could occur).

Instead you could modify the existing code in sequenza.results and trying to avoid the steps that are causing the problem (it is probably the alternative solution plot…)

Best

Francesco

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Дмитрий Голик

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Jul 6, 2021, 11:55:06 AM7/6/21

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Hello!

I ran into similar problem, and i found out that sequenza does not like chromosomes being not in reference order. I spent two days figuring. I think it is sequenza bug.

E. g. if i convert bam2seqz in parallel and then merge results as advised:

sequenza-utils.py bam2seqz --parallel 8 -C chr1 chr2 chr3 chr4 chr5 chr6 chr7 chr8 chr9 chr10 chr11 chr12 chr13 chr14 chr15 chr16 chr17 chr18 chr19 chr20 chr21 chr22 chrX chrY chrM -n normal.bam -t tumor.bam --fasta hg38.analysisSet.fa -gc hg38.gc50Base.wig.gz -o out.seqz.gz

zcat *.seqz.gz | gawk '{if (NR!=1 && $1 != "chromosome") {print $0}}' | bgzip > out.seqz.gz

, then chromosomes will follow in lexicographical order:

chr1 chr10 chr11 chr12 chr13 chr14 chr15 chr16 chr17 chr18 chr19 chr2 chr20 chr21 chr22 chr3 chr4 chr5 chr6 chr7 chr8 chr9 chrM chrX chrY

, and sequenza fit gives the error (and also the first line in the result is lost).

So now i place chromosomes in the reference order:

chr1 chr2 chr3 chr4 chr5 chr6 chr7 chr8 chr9 chr10 chr11 chr12 chr13 chr14 chr15 chr16 chr17 chr18 chr19 chr20 chr21 chr22 chrX chrY chrM

CHROMOSOMES=`awk 'BEGIN{c="chr"; for (a=1; a <= 22; a++) print c a}'`" chrX chrY chrM"

CHR_FILES=`echo $CHROMOSOMES | sed 's/\s\+/\n/g' | awk '{print "out_" $0 ".seqz.gz"}'`

zcat $CHR_FILES | gawk -v SEQ_HEADER="`zcat $CHR_FILES | head -n 1`" 'BEGIN{print SEQ_HEADER} {if (NR!=1 && $1 != "chromosome") {print $0}}' | bgzip > out.seqz.gz

tabix -f -s 1 -b 2 -e 2 -S 1 out.seqz.gz

And sequenza runs perfectly fine despite the data being basically the same!

I think at least the above script should be part of the original bam2seqz in parallel mode. And at most there should be detailed investigation, whether this bug affects validity of sequenza results even when this error does not occur. Unfortunately i can not make this investigation cos i basically have no clue what sequenza does. :'(

четверг, 30 марта 2017 г. в 04:35:00 UTC+3, amit...@gmail.com:

Дмитрий Голик

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Jul 9, 2021, 5:42:42 AM7/9/21

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I actually discovered the root of the problem (at least in my case). https://bitbucket.org/sequenzatools/sequenza/issues/19/error-in-seqdefault-finite-number-issue

1. I included chrM in my chromosome list.

2. chrM is only about 17k long.

3. windowValues function can not properly handle such short intervals so it gives NA in chrM_ratio$chrM$end (and maybe there are other similar errors but this error comes first).

4. sequenza.results processes only 24 first chromosomes by default.

5. So when my chromosomes come in lexicographical order, it takes chrM, and when they are in reference order, chrM is 25th chromosome so it is ignored.

So basically it means that sequenza was not made to process chrM, and it should not be given to sequenza input. However i think sequenza should warn me about this rather than telling me magical curse words.

вторник, 6 июля 2021 г. в 18:55:06 UTC+3, Дмитрий Голик:

Message has been deleted

Duo Xie

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Sep 29, 2022, 4:18:46 AM9/29/22

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My suggestion to solve this issue is to filter the chromosomes with NA:

CP.tumor.extr <- sequenza.fit(tumor.extr,chromosome.list = c(paste0("chr", c(1:22))) )
for(i in names(tumor.extr$segments)){
    dfm <- tumor.extr$segments[[i]]
    rownum <- nrow(dfm[which(!is.na(dfm$Bf)),])
    if(rownum>0){chrlist <- c(chrlist,i)}
}
sequenza.results(sequenza.extract = tumor.extr,cp.table = CP.tumor.extr, sample.id = outname,out.dir=outdir, chromosome.list = chrlist)

Best,

Duo

Donkiss Zidane

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Donkiss Zidane

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