Indexing the genome

[sebel76]

Hi Atul,

In response to your email of yesterday. I weird, it works on your computer… Yes, I used ‘python3’ and bowtie ‘v1’.

As troubleshooting, I try some things. (1) Build the index by myself and indicate the path of the index in the ‘phaser.set’ file. It is not working. (2) I used only for the 7 chromosomes without unassembled scaffold and contig. It is not working. (3) I download a second time the genome and tried against. It is not working.

See the log information bellow:

mbpdesebastien:phasiRNA sebel76$ python3 phaser

Verifying User Authorization

Hello 'sebel76' - Please report issues at: https://github.com/atulkakrana/phasTER/issues

Fn: checkLibs

--Python v3.0 or higher : found --Perl v5.14 or higher : found --Bowtie (v1) : found --Scalar::Util (perl) : found --Data::Dumper (perl) : found --Parallel::ForkManager (perl) : found --Getopt::Long (perl) : found --phasTER Core : found

Fn: Settings Reader

User Input runType: G User Input reference location: /Users/sebel76/NucleicAcid/ensembl_relase31/dna/HvuASM32608v1Chr.fa User Input Libs: ['UNMDay0clippedCollapsed.txt'] User Input to auto fetch libs: T User Input for phase length: 21 User Input index location:

7 cores reserved for analysis

7 threads assigned to one lib

This is first run - create index

Fn: indexBuilder

Reference file located - Preparing to create index PHASER uses FASTA header as key for identifying the phased loci Cleaning header '/Users/sebel76/NucleicAcid/ensembl_relase31/dna/HvuASM32608v1Chr.fa' reference FASTA file Traceback (most recent call last): File "phaser.py", line 990, in File "phaser.py", line 854, in main File "phaser.py", line 572, in indexBuilder File "phaser.py", line 653, in FASTAClean OSError: [Errno 22] Invalid argument

Have you an idea how I can resolve the problem? Thanks, Sébastien

[atulkakrana]

Sebastian,

The issue is not related to indexing. Phaser is trying to check your FASTA file, and clean messy headers, if any. It seems its not being able to locate your FASTA file. Which operating system are you using? and could you please post contents of your settings file (phaser.set), only the settings portion of it?

Atul

[atulkakrana]

I believe phasTER binaries are in the same folder as your reference FASTA and sRNA library. If that is true, then can you provide reference path like this (in settings file):

@reference = ./Hordeum_vulgare.ASM32608v1.fa

Sometimes the $PATH variable is set differently (or incorrectly) on Linux systems, and do not represent true location of a file. Providing the path, as I included above should fix the problem in such cases.

Atul

[sebel76]

Hello Atul,

This is the answer of your first post: I work with a Unix operating system on a mac os x 10.12.2. See the contents of my settings file (phaser.set):

<<< Settings file for phaser script >>>

<<< Mandatory Settings, value in text, see descriptions below >>> @runType = G @reference = /Users/sebel76/NucleicAcid/ensembl_relase31/dna/HvuASM32608v1Chr.fa @userLibs = UNMDay0clippedCollapsed.txt @libFormat = T
@phase = 21

Thanks, Sébastien

[sebel76]

Hello Atul,

This is the answer of your second post: I tried to change the folder of the reference genome fasta file and put this one in the same folder of the phasTER binaries sRNA library. I use you recommandation by changing the setting file:

@reference = ./Hordeum_vulgare.ASM32608v1.fa

See the contents of my settings file (phaser.set):

<<< Settings file for phaser script >>>

<<< Mandatory Settings, value in text, see descriptions below >>> @runType = G @reference = ./HvuASM32608v1Chr.fa @userLibs = UNMDay0clippedCollapsed.txt @libFormat = T
@phase = 21

Then, I run the phaser script and I receive the same same message error:

mbpdesebastien:phasiRNA sebel76$ python3 phaser

Verifying User Authorization

Hello 'sebel76' - Please report issues at: https://github.com/atulkakrana/phasTER/issues

Fn: checkLibs

--Python v3.0 or higher : found --Perl v5.14 or higher : found --Bowtie (v1) : found --Scalar::Util (perl) : found --Data::Dumper (perl) : found --Parallel::ForkManager (perl) : found --Getopt::Long (perl) : found --phasTER Core : found

Fn: Settings Reader

User Input runType: G User Input reference location: ./HvuASM32608v1Chr.fa User Input Libs: ['UNMDay0clippedCollapsed.txt'] User Input to auto fetch libs: T User Input for phase length: 21 User Input index location:

7 cores reserved for analysis

7 threads assigned to one lib

This is first run - create index

Fn: indexBuilder

Reference file located - Preparing to create index PHASER uses FASTA header as key for identifying the phased loci Cleaning header './HvuASM32608v1Chr.fa' reference FASTA file Traceback (most recent call last): File "phaser.py", line 990, in File "phaser.py", line 854, in main File "phaser.py", line 572, in indexBuilder File "phaser.py", line 653, in FASTAClean OSError: [Errno 22] Invalid argument mbpdesebastien:phasiRNA sebel76$

I don't understand... Thinks you that is the reference file header that create the problem?

Thanks, Sébastien

[atulkakrana]

This is very strange, phasTER is not being able to find the file !! I must say that we never test software on MacOS and it's pretty common. But since it's Unix the scripts should just work fine. Unfortunately, I can't test it on Mac.

Please send me the list of files from you analysis folder. Just navigate to folder where you have binaries and reference FASTA and execute ls -lah. Send me the output. Also, please send me the output of pwd command, execute it from same folder.

Atul

[sebel76]

Sorry for the delay of response...

See bellow the information requested:

MacBook-Pro-de-Sebastien:phasiRNA sebel76$ ls -lah total 8000880 drwxr-xr-x 14 sebel76 staff 476B 20 fév 14:27 . drwxr-xr-x 15 sebel76 staff 510B 20 fév 14:26 .. -rw-r--r--@ 1 sebel76 staff 6,0K 20 fév 14:26 .DS_Store -rw-r--r-- 1 sebel76 staff 52K 26 déc 20:19 .Rhistory -rw-r--r-- 1 sebel76 staff 0B 20 fév 14:27 HvuASM32608v1Chr.clean.fa -rw-r--r--@ 1 sebel76 staff 3,7G 13 fév 18:59 HvuASM32608v1Chr.fa -rw-r--r-- 1 sebel76 staff 9B 20 fév 14:27 HvuASM32608v1Chr.summ.txt -rw-r--r-- 1 sebel76 staff 122M 8 fév 11:13 UNMDay0clippedCollapsedTagCount.txt -rwxr-xr-x@ 1 sebel76 staff 821K 7 mai 2012 bowtie -rwxr--r-- 1 sebel76 staff 42K 7 fév 09:43 collapser -rwxr--r-- 1 sebel76 staff 23K 7 fév 09:43 phaser -rw-r--r--@ 1 sebel76 staff 1,2K 13 fév 10:27 phaser.set -rwxr--r-- 1 sebel76 staff 37K 7 fév 09:43 revferno -rw-r--r-- 1 sebel76 staff 0B 20 fév 14:27 runtime_02_20_14_27

MacBook-Pro-de-Sebastien:phasiRNA sebel76$ pwd /Users/sebel76/paper/4/phasiRNA

Thanks for your help! Sébastien