[zillur]

Thank you very much. I am a neophyte here and making mistakes in my every steps but you are always helping me. Its amazing for me. Thank you so much. However I have got my terminal back. I had to type
/bin/echo 'export PATH=${PATH}:/usr/bin:/bin:/usr/sbin:/sbin:/usr/local/bin:/opt/X11/bin' >> ~/.bash_profile then hit enter. Now its working
But now my bowtie2 command is not working. Should I create path using vi editor?
Mds-MacBook-Pro:desktop ZILLURRAHMAN$ type ls
ls is hashed (/bin/ls)
Mds-MacBook-Pro:desktop ZILLURRAHMAN$ type bowtie2
-bash: type: bowtie2: not found
Mds-MacBook-Pro:desktop ZILLURRAHMAN$

Should I create path using vi editor?

Mds-MacBook-Pro:desktop ZILLURRAHMAN$ cd bioinformatics/bow-tie/bowtie2-2.2.3/
Mds-MacBook-Pro:bowtie2-2.2.3 ZILLURRAHMAN$ ls
AUTHORS bowtie2-align-l-debug bowtie2-inspect
LICENSE bowtie2-align-s bowtie2-inspect-l
MANUAL bowtie2-align-s-debug bowtie2-inspect-l-debug
MANUAL.markdown bowtie2-build bowtie2-inspect-s
NEWS bowtie2-build-l bowtie2-inspect-s-debug
TUTORIAL bowtie2-build-l-debug doc
VERSION bowtie2-build-s example
bowtie2 bowtie2-build-s-debug scripts
bowtie2-align-l bowtie2-buildc
Mds-MacBook-Pro:bowtie2-2.2.3 ZILLURRAHMAN$ pwd
/Users/ZILLURRAHMAN/desktop/bioinformatics/bow-tie/bowtie2-2.2.3
Mds-MacBook-Pro:bowtie2-2.2.3 ZILLURRAHMAN$
Should I type:
export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/Bow-tie/bowtie2-2.2.3/:$PATH"
then save and quit the vi editor? I am asking you mentioning my details files and folders cause I don't another mess. Hope you understand my situation here.

Best Regards
Zillur

[GenoMax]

export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/[COLOR="Red"]b[/COLOR]ow-tie/bowtie2-2.2.3/:$PATH"

Make sure you use the correct case (you had a capital B in the line I copied above". Just do that on the command line before you go editing any files. OS should now be able to find bowtie2.

[zillur]

Thank you very much. So I should just need to type it in the command line. Don't I need any vi editor?

[GenoMax]

You can add that line to your .bash_profile using vi after you test it on the command line and confirm that it works. Then it would be automatically sourced when you log in next time since it will be there in the profile.

[zillur]

Thank you very much. But I can't do it using vi editor. I don't know what is happening here. But I just typed the line in the terminal and its working now. But is it safe to work direct with terminal?
Mds-MacBook-Pro:~ ZILLURRAHMAN$ type bowtie2
bowtie2 is hashed (/Users/ZILLURRAHMAN/Desktop/Bioinformatics/bow-tie/bowtie2-2.2.3/bowtie2)

Using same trick can I make path for samtools?

[GenoMax]

It is safe to work in the terminal. Adding the path modifier to profile file is a convenience so you don't have to do it in the terminal each time. You can use the same method to add path for samtools (and any additional programs you need).

[zillur]

Thank you very much. Its really great to find some people like you. Now I am able to create paths for programs that I need.

Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ pwd
/Users/ZILLURRAHMAN/desktop/bioinformatics/samtools/samtools-1.1
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/samtools/samtools-1.1/:$PATH"
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ type samtools.1
samtools.1 is /Users/ZILLURRAHMAN/Desktop/Bioinformatics/samtools/samtools-1.1/samtools.1
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ type bowtie2
bowtie2 is /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bow-tie/bowtie2-2.2.3/bowtie2
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$

But another problem, every time I close the terminal it loses all the newly created paths. Even if I open a new tab on the same terminal window it can't find the newly created paths.

Last login: Sun Nov 2 17:00:11 on ttys000
-bash: $: command not found
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ type bowtie2
-bash: type: bowtie2: not found
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$ type samtools.1
-bash: type: samtools.1: not found
Mds-MacBook-Pro:samtools-1.1 ZILLURRAHMAN$

What should I do now? I just don't want to create path every time I am opening a new shell.

Best Regards
Zillur

[GenoMax]

This is a simple work around.

Since you are having trouble editing .bash_profile file put the PATH command in a text file (let us call the file "set_path_file") using an editor. Then when you open a new terminal do the following

$ source set_path_file

That should set the PATH variable correctly for that terminal session. You will need to do this for each new terminal you open.

[zillur]

Thank you very much. I am working on this. Meanwhile I have another problem.

Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.fa lambdaout.rev.1.bt2
eg2.sam lambda_virus.rev.1.bt2 lambdaout.rev.2.bt2
egl.sam lambda_virus.rev.2.bt2 nohup.out
lambda_virus.1.bt2 lambdaout.1.bt2 out
lambda_virus.2.bt2 lambdaout.2.bt2 out.sam
lambda_virus.3.bt2 lambdaout.3.bt2
lambda_virus.4.bt2 lambdaout.4.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools view -bS eg2.sam > eg2.bam
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.4.bt2 lambdaout.4.bt2
eg2.bam lambda_virus.fa lambdaout.rev.1.bt2
eg2.sam lambda_virus.rev.1.bt2 lambdaout.rev.2.bt2
egl.sam lambda_virus.rev.2.bt2 nohup.out
lambda_virus.1.bt2 lambdaout.1.bt2 out
lambda_virus.2.bt2 lambdaout.2.bt2 out.sam
lambda_virus.3.bt2 lambdaout.3.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$ more eg2.bam
"eg2.bam" may be a binary file. See it anyway?
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools sort eg2.bam eg2.sorted
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.3.bt2 lambdaout.3.bt2
eg2.bam lambda_virus.4.bt2 lambdaout.4.bt2
eg2.sam lambda_virus.fa lambdaout.rev.1.bt2
eg2.sorted.bam lambda_virus.rev.1.bt2 lambdaout.rev.2.bt2
egl.sam lambda_virus.rev.2.bt2 nohup.out
lambda_virus.1.bt2 lambdaout.1.bt2 out
lambda_virus.2.bt2 lambdaout.2.bt2 out.sam
Mds-MacBook-Pro:reference ZILLURRAHMAN$ more eg2.sorted.bam
"eg2.sorted.bam" may be a binary file. See it anyway?
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools mpileup -uf /example/reference/lambda_virus.fa eg2.sorted.bam | bcftools view -bvcg - > eg2.raw.bcf
[fai_load] build FASTA index.
[fai_build] fail to open the FASTA file /example/reference/lambda_virus.fa
[fai_load] fail to open FASTA index.
-bash: bcftools: command not found
Mds-MacBook-Pro:reference ZILLURRAHMAN$

From where I can get the "bcftools" files?

[GenoMax]

From where I can get the "bcftools" files?

You have to download and compile bcftools from here: http://sourceforge.net/projects/samt...r.bz2/download

[zillur]

Thank you very much. I downloaded and compiled it using "make" command. Then tried to create path for "bcftools" using previous trick but it is not working here.

Mds-MacBook-Pro:bioinformatics ZILLURRAHMAN$ cd bcftools-1.1/
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ ls
AUTHORS em.c prob1.c vcfconcat.c vcfquery.c
HMM.c em.o prob1.h vcfconcat.o vcfquery.o
HMM.h filter.c prob1.o vcfconvert.c vcfroh.c
HMM.o filter.h rbuf.h vcfconvert.o vcfroh.o
INSTALL filter.o reheader.c vcffilter.c vcfsom.c
LICENSE gvcf.c reheader.o vcffilter.o vcfsom.o
Makefile gvcf.o tabix.c vcfgtcheck.c vcfstats.c
README htslib-1.1 tabix.o vcfgtcheck.o vcfstats.o
bcftools khash_str2str.h test vcfindex.c vcfutils.pl
bcftools.1 kmin.c tsv2vcf.c vcfindex.o vcfview.c
bcftools.h kmin.h tsv2vcf.h vcfisec.c vcfview.o
bcftools.txt kmin.o tsv2vcf.o vcfisec.o vcmp.c
call.h main.c vcfannotate.c vcfmerge.c vcmp.h
ccall.c main.o vcfannotate.o vcfmerge.o vcmp.o
ccall.o mcall.c vcfcall.c vcfnorm.c version.c
convert.c mcall.o vcfcall.o vcfnorm.o version.h
convert.h plot-vcfstats vcfcnv.c vcfplugin.c
convert.o plugins vcfcnv.o vcfplugin.o
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ pwd
/Users/ZILLURRAHMAN/desktop/bioinformatics/bcftools-1.1
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools/:$PATH"
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ type bcftools
-bash: type: bcftools: not found
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$

Would you please to tell me which point I missed?

Best regards
Zillur

[GenoMax]

You need to add execute permissions to the new bcftools executable.

$ chmod u+x /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools/bcftools

[zillur]

Thank you very much. May be I am missing something. Still bcftools command can't be found.

Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ chmod u+x /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools/bcftools
chmod: /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools/bcftools: Not a directory
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ chmod u+x /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ type bcftools
-bash: type: bcftools: not found
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools/:$PATH"
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ type bcftools
-bash: type: bcftools: not found
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$

[GenoMax]

You added an extra bcftools word at the end of your path command. Command should be

$ export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1:$PATH"

[zillur]

Thank you very much. Its working now. But......


Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ export PATH="/Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/:$PATH"
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ type bcftools
bcftools is /Users/ZILLURRAHMAN/Desktop/Bioinformatics/bcftools-1.1/bcftools
Mds-MacBook-Pro:bcftools-1.1 ZILLURRAHMAN$ cd ..
Mds-MacBook-Pro:bioinformatics ZILLURRAHMAN$ cd bow-tie/bowtie2-2.2.3/example/reference/
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.3.bt2 lambdaout.4.bt2
eg2.bam lambda_virus.4.bt2 lambdaout.rev.1.bt2
eg2.raw.bcf lambda_virus.fa lambdaout.rev.2.bt2
eg2.sam lambda_virus.rev.1.bt2 nohup.out
eg2.sorted.bam lambda_virus.rev.2.bt2 out
egl.sam lambdaout.1.bt2 out.sam
lambda_virus.1.bt2 lambdaout.2.bt2
lambda_virus.2.bt2 lambdaout.3.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools mpileup -uf /example/reference/lambda_virus.fa eg2.sorted.bam | bcftools view -bvcg - > eg2.raw.bcf
bcftools: invalid option -- b

About: VCF/BCF conversion, view, subset and filter VCF/BCF files.
Usage: bcftools view [options] <in.vcf.gz> [region1 [...]]

Output options:
-G, --drop-genotypes drop individual genotype information (after subsetting if -s option set)
-h/H, --header-only/--no-header print the header only/suppress the header in VCF output
-l, --compression-level [0-9] compression level: 0 uncompressed, 1 best speed, 9 best compression [-1]
-o, --output-file <file> output file name [stdout]
-O, --output-type <b|u|z|v> b: compressed BCF, u: uncompressed BCF, z: compressed VCF, v: uncompressed VCF [v]
-r, --regions <region> restrict to comma-separated list of regions
-R, --regions-file <file> restrict to regions listed in a file
-t, --targets [^]<region> similar to -r but streams rather than index-jumps. Exclude regions with "^" prefix
-T, --targets-file [^]<file> similar to -R but streams rather than index-jumps. Exclude regions with "^" prefix

Subset options:
-a, --trim-alt-alleles trim alternate alleles not seen in the subset
-I, --no-update do not (re)calculate INFO fields for the subset (currently INFO/AC and INFO/AN)
-s, --samples [^]<list> comma separated list of samples to include (or exclude with "^" prefix)
-S, --samples-file [^]<file> file of samples to include (or exclude with "^" prefix)
--force-samples only warn about unknown subset samples

Filter options:
-c/C, --min-ac/--max-ac <int>[:<type>] minimum/maximum count for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-f, --apply-filters <list> require at least one of the listed FILTER strings (e.g. "PASS,.")
-g, --genotype [^]<hom|het|miss> require one or more hom/het/missing genotype or, if prefixed with "^", exclude sites with hom/het/missing genotypes
-i/e, --include/--exclude <expr> select/exclude sites for which the expression is true (see man page for details)
-k/n, --known/--novel select known/novel sites only (ID is not/is '.')
-m/M, --min-alleles/--max-alleles <int> minimum/maximum number of alleles listed in REF and ALT (e.g. -m2 -M2 for biallelic sites)
-p/P, --phased/--exclude-phased select/exclude sites where all samples are phased
-q/Q, --min-af/--max-af <float>[:<type>] minimum/maximum frequency for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-u/U, --uncalled/--exclude-uncalled select/exclude sites without a called genotype
-v/V, --types/--exclude-types <list> select/exclude comma-separated list of variant types: snps,indels,mnps,other [null]
-x/X, --private/--exclude-private select/exclude sites where the non-reference alleles are exclusive (private) to the subset samples

[fai_load] build FASTA index.
[fai_build] fail to open the FASTA file /example/reference/lambda_virus.fa
[fai_load] fail to open FASTA index.
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.3.bt2 lambdaout.4.bt2
eg2.bam lambda_virus.4.bt2 lambdaout.rev.1.bt2
eg2.raw.bcf lambda_virus.fa lambdaout.rev.2.bt2
eg2.sam lambda_virus.rev.1.bt2 nohup.out
eg2.sorted.bam lambda_virus.rev.2.bt2 out
egl.sam lambdaout.1.bt2 out.sam
lambda_virus.1.bt2 lambdaout.2.bt2
lambda_virus.2.bt2 lambdaout.3.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools mpileup -uf /lambda_virus.fa eg2.sorted.bam | bcftools view -bvcg - > eg2.raw.bcf
bcftools: invalid option -- b

About: VCF/BCF conversion, view, subset and filter VCF/BCF files.
Usage: bcftools view [options] <in.vcf.gz> [region1 [...]]

Output options:
-G, --drop-genotypes drop individual genotype information (after subsetting if -s option set)
-h/H, --header-only/--no-header print the header only/suppress the header in VCF output
-l, --compression-level [0-9] compression level: 0 uncompressed, 1 best speed, 9 best compression [-1]
-o, --output-file <file> output file name [stdout]
-O, --output-type <b|u|z|v> b: compressed BCF, u: uncompressed BCF, z: compressed VCF, v: uncompressed VCF [v]
-r, --regions <region> restrict to comma-separated list of regions
-R, --regions-file <file> restrict to regions listed in a file
-t, --targets [^]<region> similar to -r but streams rather than index-jumps. Exclude regions with "^" prefix
-T, --targets-file [^]<file> similar to -R but streams rather than index-jumps. Exclude regions with "^" prefix

Subset options:
-a, --trim-alt-alleles trim alternate alleles not seen in the subset
-I, --no-update do not (re)calculate INFO fields for the subset (currently INFO/AC and INFO/AN)
-s, --samples [^]<list> comma separated list of samples to include (or exclude with "^" prefix)
-S, --samples-file [^]<file> file of samples to include (or exclude with "^" prefix)
--force-samples only warn about unknown subset samples

Filter options:
-c/C, --min-ac/--max-ac <int>[:<type>] minimum/maximum count for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-f, --apply-filters <list> require at least one of the listed FILTER strings (e.g. "PASS,.")
-g, --genotype [^]<hom|het|miss> require one or more hom/het/missing genotype or, if prefixed with "^", exclude sites with hom/het/missing genotypes
-i/e, --include/--exclude <expr> select/exclude sites for which the expression is true (see man page for details)
-k/n, --known/--novel select known/novel sites only (ID is not/is '.')
-m/M, --min-alleles/--max-alleles <int> minimum/maximum number of alleles listed in REF and ALT (e.g. -m2 -M2 for biallelic sites)
-p/P, --phased/--exclude-phased select/exclude sites where all samples are phased
-q/Q, --min-af/--max-af <float>[:<type>] minimum/maximum frequency for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-u/U, --uncalled/--exclude-uncalled select/exclude sites without a called genotype
-v/V, --types/--exclude-types <list> select/exclude comma-separated list of variant types: snps,indels,mnps,other [null]
-x/X, --private/--exclude-private select/exclude sites where the non-reference alleles are exclusive (private) to the subset samples

[fai_load] build FASTA index.
[fai_build] fail to open the FASTA file /lambda_virus.fa
[fai_load] fail to open FASTA index.
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.3.bt2 lambdaout.4.bt2
eg2.bam lambda_virus.4.bt2 lambdaout.rev.1.bt2
eg2.raw.bcf lambda_virus.fa lambdaout.rev.2.bt2
eg2.sam lambda_virus.rev.1.bt2 nohup.out
eg2.sorted.bam lambda_virus.rev.2.bt2 out
egl.sam lambdaout.1.bt2 out.sam
lambda_virus.1.bt2 lambdaout.2.bt2
lambda_virus.2.bt2 lambdaout.3.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$ samtools mpileup -uf /reference/lambda_virus.fa eg2.sorted.bam | bcftools view -bvcg - > eg2.raw.bcf
bcftools: invalid option -- b

About: VCF/BCF conversion, view, subset and filter VCF/BCF files.
Usage: bcftools view [options] <in.vcf.gz> [region1 [...]]

Output options:
-G, --drop-genotypes drop individual genotype information (after subsetting if -s option set)
-h/H, --header-only/--no-header print the header only/suppress the header in VCF output
-l, --compression-level [0-9] compression level: 0 uncompressed, 1 best speed, 9 best compression [-1]
-o, --output-file <file> output file name [stdout]
-O, --output-type <b|u|z|v> b: compressed BCF, u: uncompressed BCF, z: compressed VCF, v: uncompressed VCF [v]
-r, --regions <region> restrict to comma-separated list of regions
-R, --regions-file <file> restrict to regions listed in a file
-t, --targets [^]<region> similar to -r but streams rather than index-jumps. Exclude regions with "^" prefix
-T, --targets-file [^]<file> similar to -R but streams rather than index-jumps. Exclude regions with "^" prefix

Subset options:
-a, --trim-alt-alleles trim alternate alleles not seen in the subset
-I, --no-update do not (re)calculate INFO fields for the subset (currently INFO/AC and INFO/AN)
-s, --samples [^]<list> comma separated list of samples to include (or exclude with "^" prefix)
-S, --samples-file [^]<file> file of samples to include (or exclude with "^" prefix)
--force-samples only warn about unknown subset samples

Filter options:
-c/C, --min-ac/--max-ac <int>[:<type>] minimum/maximum count for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-f, --apply-filters <list> require at least one of the listed FILTER strings (e.g. "PASS,.")
-g, --genotype [^]<hom|het|miss> require one or more hom/het/missing genotype or, if prefixed with "^", exclude sites with hom/het/missing genotypes
-i/e, --include/--exclude <expr> select/exclude sites for which the expression is true (see man page for details)
-k/n, --known/--novel select known/novel sites only (ID is not/is '.')
-m/M, --min-alleles/--max-alleles <int> minimum/maximum number of alleles listed in REF and ALT (e.g. -m2 -M2 for biallelic sites)
-p/P, --phased/--exclude-phased select/exclude sites where all samples are phased
-q/Q, --min-af/--max-af <float>[:<type>] minimum/maximum frequency for non-reference (nref), 1st alternate (alt1) or minor (minor) alleles [nref]
-u/U, --uncalled/--exclude-uncalled select/exclude sites without a called genotype
-v/V, --types/--exclude-types <list> select/exclude comma-separated list of variant types: snps,indels,mnps,other [null]
-x/X, --private/--exclude-private select/exclude sites where the non-reference alleles are exclusive (private) to the subset samples

[fai_load] build FASTA index.
[fai_build] fail to open the FASTA file /reference/lambda_virus.fa
[fai_load] fail to open FASTA index.
Mds-MacBook-Pro:reference ZILLURRAHMAN$ ls
commands lambda_virus.3.bt2 lambdaout.4.bt2
eg2.bam lambda_virus.4.bt2 lambdaout.rev.1.bt2
eg2.raw.bcf lambda_virus.fa lambdaout.rev.2.bt2
eg2.sam lambda_virus.rev.1.bt2 nohup.out
eg2.sorted.bam lambda_virus.rev.2.bt2 out
egl.sam lambdaout.1.bt2 out.sam
lambda_virus.1.bt2 lambdaout.2.bt2
lambda_virus.2.bt2 lambdaout.3.bt2
Mds-MacBook-Pro:reference ZILLURRAHMAN$

[fai_load] build FASTA index.
[fai_build] fail to open the FASTA file /reference/lambda_virus.fa
[fai_load] fail to open FASTA index.

What does it mean?

Best Regards
Zillur