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ZW-xjtlu / exomePeak2

Peak calling and differential methylation for MeRIP-Seq
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an error occured #1

Open sunhaifeng123 opened 5 years ago

sunhaifeng123 commented 5 years ago

Hi

exomePeak2 is so nice a better package for m6a , I installed it correctly and try to use , all bam files have been indexed. I come across the error like this :

My command is :

library(exomePeak2)
library(TxDb.Mmusculus.UCSC.mm10.ensGene)
library(BSgenome.Mmusculus.UCSC.mm10)

MeRIP_Seq_Alignment <- scanMeripBAM(
  bam_ip = c("R19008728-QMJ-Thy--control-IP-1_combined_R.sorted.bam", 
             "R19008728-QMJ-Thy--control-IP-2_combined_R.sorted.bam"), 
  bam_input = c("R19008728-QMJ-Thy--control-input-1_combined_R.sorted.bam",
                "R19008728-QMJ-Thy--control-input-2_combined_R.sorted.bam"),
  bam_treated_ip = c("R19008728-QMJ-Thy--KO-IP-1_combined_R.sorted.bam", 
                     "R19008728-QMJ-Thy--KO-IP-2_combined_R.sorted.bam"),
  bam_treated_input = c("R19008728-QMJ-Thy--KO-input-1_combined_R.sorted.bam",
                        "R19008728-QMJ-Thy--KO-input-2_combined_R.sorted.bam"), 
  paired_end = TRUE,
  library_type = "1st_strand") 

SummarizedExomePeaks <- exomePeakCalling(merip_bams = MeRIP_Seq_Alignment,
                                         txdb = TxDb.Mmusculus.UCSC.mm10.ensGene,
                                         bsgenome = Mmusculus,
                                         p_adj_cutoff = 0.05,
                                         logFC_cutoff = 0,
                                         parallel = TRUE)

And then it runs and error : 

Generating bins on exons...
Counting reads on bins...
Error in names(res) <- nms : 'names'属性的长度[8]必需和矢量的长度[2]一样

> Error in serialize(data, node$con, xdr = FALSE) : ignoring SIGPIPE signal

I'll very grateful if you can give me any help , thank you !

Sun Haifeng NanJing Medical University

sunhaifeng123 commented 5 years ago

Hi

It seems I use the parameter ： parallel = TRUE which result in my PC's memory out

After remove it everything seems run well , except another errored in the end :

Error in makeGuitarCoordsFromTxDb(txdb) :
  没有"makeGuitarCoordsFromTxDb"这个函数

I have installed the R package "Guitar" and my R version is 3.6.0

Thanks in advance if you can reply to me about this problem

Best

Sun Haifeng 20190517 NanJing Medical University

ZW-xjtlu commented 5 years ago

Dear Haifeng,

The error of guitar is now fixed, your analysis will be success after re-install the package exomePeak2 with install_github(). For the memory out issue, I reduced the default number of parallels used if user set parallel = TRUE, your PC may able to run in parallel at this time.

Sincerely, Zhen

在 2019年5月17日，下午1:15，sunhaifeng123 notifications@github.com 写道：

Hi

It seems I use the parameter ： parallel = TRUE which result in my PC's memory out

After remove it everything seems run well , except another errored in the end :

Error in makeGuitarCoordsFromTxDb(txdb) : 没有"makeGuitarCoordsFromTxDb"这个函数 I have installed the R package "Guitar" and my R version is 3.6.0

Thanks in advance if you can reply to me about this problem

Best

Sun Haifeng 20190517 NanJing Medical University

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ZW-xjtlu commented 5 years ago

Dear Haifeng,

It seems that the error message comes from the function summarizeOverlaps() in the bioconductor package GenomicAlignments. So, please first check the sizes of your bam files on disk, make sure they are not broken and normal in sizes. You may also try to re-install exomePeak2 package with install_github() again, I test the same code of yours using my own bam files, and it is working.

Best Wishes, Zhen

在 2019年5月15日，下午10:53，sunhaifeng123 notifications@github.com 写道：

Hi

exomePeak2 is so nice a better package for m6a , I installed it correctly and try to use , all bam files have been indexed. I come across the error like this :

My command is :

library(exomePeak2) library(TxDb.Mmusculus.UCSC.mm10.ensGene) library(BSgenome.Mmusculus.UCSC.mm10)

MeRIP_Seq_Alignment <- scanMeripBAM( bam_ip = c("R19008728-QMJ-Thy--control-IP-1_combined_R.sorted.bam", "R19008728-QMJ-Thy--control-IP-2_combined_R.sorted.bam"), bam_input = c("R19008728-QMJ-Thy--control-input-1_combined_R.sorted.bam", "R19008728-QMJ-Thy--control-input-2_combined_R.sorted.bam"), bam_treated_ip = c("R19008728-QMJ-Thy--KO-IP-1_combined_R.sorted.bam", "R19008728-QMJ-Thy--KO-IP-2_combined_R.sorted.bam"), bam_treated_input = c("R19008728-QMJ-Thy--KO-input-1_combined_R.sorted.bam", "R19008728-QMJ-Thy--KO-input-2_combined_R.sorted.bam"), paired_end = TRUE, library_type = "1st_strand")

SummarizedExomePeaks <- exomePeakCalling(merip_bams = MeRIP_Seq_Alignment, txdb = TxDb.Mmusculus.UCSC.mm10.ensGene, bsgenome = Mmusculus, p_adj_cutoff = 0.05, logFC_cutoff = 0, parallel = TRUE) And then it runs and error :

Generating bins on exons... Counting reads on bins... Error in names(res) <- nms : 'names'属性的长度[8]必需和矢量的长度[2]一样

Error in serialize(data, node$con, xdr = FALSE) : ignoring SIGPIPE signal I'll very grateful if you can give me any help , thank you !

Sun Haifeng NanJing Medical University

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sunhaifeng123 commented 5 years ago

Hi ZhenWei,

Thank you very much for reply , I reinstalled exomePeak2 and use well except a parameter "save_plot_analysis = TRUE" , to be exact , I think it's Guitar's problem , because when I run : 

> plotGuitar(SummarizedExomePeaks, txdb = TxDb.Mmusculus.UCSC.mm10.ensGene)
错误: 'makeGuitarCoordsFromTxDb' is not an exported object from 'namespace:Guitar'

Maybe I'm using Ubuntu18 ? 

> sessionInfo()
R version 3.6.0 (2019-04-26)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 18.04.2 LTS

Matrix products: default
BLAS:   /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.7.1
LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.7.1

locale:
 [1] LC_CTYPE=zh_CN.UTF-8       LC_NUMERIC=C
 [3] LC_TIME=zh_CN.UTF-8        LC_COLLATE=zh_CN.UTF-8
 [5] LC_MONETARY=zh_CN.UTF-8    LC_MESSAGES=zh_CN.UTF-8
 [7] LC_PAPER=zh_CN.UTF-8       LC_NAME=C
 [9] LC_ADDRESS=C               LC_TELEPHONE=C
[11] LC_MEASUREMENT=zh_CN.UTF-8 LC_IDENTIFICATION=C

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base

loaded via a namespace (and not attached):
[1] compiler_3.6.0

looking for your reply

Best,

Haifeng

ZW-xjtlu commented 5 years ago

Dear Haifeng,

The error message means that the Guitar package installed on your computer does not export the function makeGuitarCoordsFromTxDb().

This error is not expected, because the Guitar package do export this function under versions >= 1.20.1.

Please update the Guitar package on your computer from bioconductor, and please check the package version of Guitar with:

packageVersion("Guitar")

Best regards, Zhen

在 2019年6月3日，下午7:03，sunhaifeng123 notifications@github.com 写道：

makeGuitarCoordsFromTxDb

sunhaifeng123 commented 5 years ago

Dear ZhenWei,

my Guitar version is 2.0.0 and i think it's the newest version ,see : https://bioconductor.org/packages/release/bioc/html/Guitar.html

> packageVersion("Guitar")
[1] ‘2.0.0’

and I don't find makeGuitarCoordsFromTxDb() from : https://bioconductor.org/packages/release/bioc/vignettes/Guitar/inst/doc/Guitar-Overview.R

Maybe it should be : makeGuitarTxdb ? because I see his scripts and Manual are modified on 26 April, 2019

Haifeng

ZW-xjtlu commented 5 years ago

Hi Haifeng,

You are right, my function name is the old one. Sorry about the mistake, I have changed it to the new one. It should be working now after you re-install exomePeak2.

Sincerely, Zhen

在 2019年6月3日，下午10:22，sunhaifeng123 notifications@github.com 写道：

makeGuitarTxdb

charishelen123 commented 4 years ago

Hi Zhenwei, I have done some m6A analysis by exomePeak2 with the following code: library(exomePeak2) GENE_ANNO_GTF = file.path("/home/ligang/m6A/exomePeak2/bam_files", "rn6.ensGene.gtf")

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.m6A.bam") IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.input.bam") INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE)

5.2 Differential Modification Analysis

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.m6A.bam") TREATED_IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.input.bam") TREATED_INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, bam_treated_input = TREATED_INPUT_BAM, bam_treated_ip = TREATED_IP_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE) but there were some warning messages as followed: Warning messages: 1: In .get_cds_IDX(mcols0$type, mcols0$phase) : The "phase" metadata column contains non-NA values for features of type stop_codon. This information was ignored. 2: In .Seqinfo.mergexy(x, y) : Each of the 2 combined objects has sequence levels not in the other:

the gtf file was downloaded from UCSC, I wanna be sure whether this analysis result is correct. looking for your reply!

Gang

ZW-xjtlu commented 4 years ago

Hi Gang,

The first warning message indicates that it ignored un expected information in the phase column of the GTF file, but it does not cause difference in the output since exomePeak2 do not use the metadata information. The second warning message arrises because the sequence levels of the GENE_ANNO_GTF file and the BAM file are inconsistent. This message will also not changing the exomePeak2 outputs because the functions in genomicRanges can automatically merge different sequence levels in different Granges objects, so no conflicts will be preserved in the downstream analysis.

Best wishes,

Zhen

在 2020年10月16日，下午10:23，charishelen123 notifications@github.com 写道：

 Hi Zhenwei, I have done some m6A analysis by exomePeak2 with the following code: library(exomePeak2) GENE_ANNO_GTF = file.path("/home/ligang/m6A/exomePeak2/bam_files", "rn6.ensGene.gtf")

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.m6A.bam") IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.input.bam") INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE)

5.2 Differential Modification Analysis

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.m6A.bam") TREATED_IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.input.bam") TREATED_INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, bam_treated_input = TREATED_INPUT_BAM, bam_treated_ip = TREATED_IP_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE) but there were some warning messages as followed: Warning messages: 1: In .get_cds_IDX(mcols0$type, mcols0$phase) : The "phase" metadata column contains non-NA values for features of type stop_codon. This information was ignored. 2: In .Seqinfo.mergexy(x, y) : Each of the 2 combined objects has sequence levels not in the other:

in 'x': chr1_AABR07046142v1_random, chr1_AABR07046150v1_random, chr1_AABR070 46158v1_random, chr1_AABR07046159v1_random, chr1_AABR07046182v1_random, chr1_AAB R07046186v1_random, chr1_AABR07046187v1_random, chr1_AABR07046211v1_random, chr1 AABR07046212v1_random, chr1_AABR07046222v1_random, chr1_AABR07046230v1_random, chr1_KL567879v1_random, chr1_KL567880v1_random, chr1_KL567881v1_random, chr1_KL5 67882v1_random, chr1_KL567883v1_random, chr1_KL567884v1_random, chr1_KL567885v1 random, chr1_KL567886v1_random, chr1_KL567887v1_random, chr1_KL567888v1_random, chr1_KL567889v1_random, chr1_KL567890v1_random, chr1_KL567893v1_random, chr1_KL5 67894v1_random, chr1_KL567895v1_random, chr2_AABR07045875v1_random, chr2_AABR070 45975v1_random, chr2_AABR07046014v1_random, chr2_KL567896v1_random, chr2_KL56789 7v1_random, chr2_KL567898v1_random, chr2_KL567899v1_random, chr2_KL567900v1_rand om, chr2_KL567901v1_random, chr2_KL567902v1_random, [... truncated] the gtf file was downloaded from UCSC, I wanna be sure whether this analysis result is correct. looking for your reply!

Gang

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charishelen123 commented 4 years ago

Thanks so much for your kind reply.

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发件人: wei lab 发送时间: 2020年10月17日 0:14 收件人: ZW-xjtlu/exomePeak2 抄送: charishelen123; Comment 主题: Re: [ZW-xjtlu/exomePeak2] an error occured (#1)

Hi Gang,

The first warning message indicates that it ignored un expected information in the phase column of the GTF file, but it does not cause difference in the output since exomePeak2 do not use the metadata information. The second warning message arrises because the sequence levels of the GENE_ANNO_GTF file and the BAM file are inconsistent. This message will also not changing the exomePeak2 outputs because the functions in genomicRanges can automatically merge different sequence levels in different Granges objects, so no conflicts will be preserved in the downstream analysis.

Best wishes,

Zhen

在 2020年10月16日，下午10:23，charishelen123 notifications@github.com 写道：

 Hi Zhenwei, I have done some m6A analysis by exomePeak2 with the following code: library(exomePeak2) GENE_ANNO_GTF = file.path("/home/ligang/m6A/exomePeak2/bam_files", "rn6.ensGene.gtf")

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.m6A.bam") IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample1.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample2.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files","Sample3.input.bam") INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE)

5.2 Differential Modification Analysis

f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.m6A.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.m6A.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.m6A.bam") TREATED_IP_BAM = c(f1,f2,f3) f1 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample4.input.bam") f2 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample5.input.bam") f3 = file.path("/home/ligang/m6A/exomePeak2/bam_files", "Sample6.input.bam") TREATED_INPUT_BAM = c(f1,f2,f3)

exomePeak2(bam_ip = IP_BAM, bam_input = INPUT_BAM, bam_treated_input = TREATED_INPUT_BAM, bam_treated_ip = TREATED_IP_BAM, gff_dir = GENE_ANNO_GTF, fragment_length = 200, genome = "rn6", paired_end = TRUE, parallel = TRUE) but there were some warning messages as followed: Warning messages: 1: In .get_cds_IDX(mcols0$type, mcols0$phase) : The "phase" metadata column contains non-NA values for features of type stop_codon. This information was ignored. 2: In .Seqinfo.mergexy(x, y) : Each of the 2 combined objects has sequence levels not in the other:

in 'x': chr1_AABR07046142v1_random, chr1_AABR07046150v1_random, chr1_AABR070 46158v1_random, chr1_AABR07046159v1_random, chr1_AABR07046182v1_random, chr1_AAB R07046186v1_random, chr1_AABR07046187v1_random, chr1_AABR07046211v1_random, chr1 AABR07046212v1_random, chr1_AABR07046222v1_random, chr1_AABR07046230v1_random, chr1_KL567879v1_random, chr1_KL567880v1_random, chr1_KL567881v1_random, chr1_KL5 67882v1_random, chr1_KL567883v1_random, chr1_KL567884v1_random, chr1_KL567885v1 random, chr1_KL567886v1_random, chr1_KL567887v1_random, chr1_KL567888v1_random, chr1_KL567889v1_random, chr1_KL567890v1_random, chr1_KL567893v1_random, chr1_KL5 67894v1_random, chr1_KL567895v1_random, chr2_AABR07045875v1_random, chr2_AABR070 45975v1_random, chr2_AABR07046014v1_random, chr2_KL567896v1_random, chr2_KL56789 7v1_random, chr2_KL567898v1_random, chr2_KL567899v1_random, chr2_KL567900v1_rand om, chr2_KL567901v1_random, chr2_KL567902v1_random, [... truncated] the gtf file was downloaded from UCSC, I wanna be sure whether this analysis result is correct. looking for your reply!

Gang

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shangguandong1996 commented 3 years ago

Dear Haifeng, The error of guitar is now fixed, your analysis will be success after re-install the package exomePeak2 with install_github(). For the memory out issue, I reduced the default number of parallels used if user set parallel = TRUE, your PC may able to run in parallel at this time. Sincerely, Zhen … 在 2019年5月17日，下午1:15，sunhaifeng123 @.***> 写道： Hi It seems I use the parameter ： parallel = TRUE which result in my PC's memory out After remove it everything seems run well , except another errored in the end : Error in makeGuitarCoordsFromTxDb(txdb) : 没有"makeGuitarCoordsFromTxDb"这个函数 I have installed the R package "Guitar" and my R version is 3.6.0 Thanks in advance if you can reply to me about this problem Best Sun Haifeng 20190517 NanJing Medical University — You are receiving this because you are subscribed to this thread. Reply to this email directly, view it on GitHub <#1>, or mute the thread https://github.com/notifications/unsubscribe-auth/AEU2CIJL6AYER7KGOMECER3PVY5QVANCNFSM4HNEEL2A.

Can I just set the core numebr by myself？ 3 threads makes work run so long ……

Li-Gaoyuan commented 3 years ago

Dear Haifeng,

I also meet the problem mentioned above that "could not find function "makeGuitarCoordsFromTxDb"", 图片 And I have the packages version as ↓ 图片

packageVersion("Trumpet") [1] ‘0.3.6’ packageVersion("Guitar") [1] ‘2.9.0’ packageVersion("exomePeak") [1] ‘2.16.0’ packageVersion("exomePeak2") [1] ‘1.5.0’

I hope to receive your suggestions, how can I run the program smoothly? Thank you.

Best Regards.