Help with Cufflinks in command line

[ccard28]

Hello,
I am new to programming/command line and am having trouble with the simplest of tasks I am sure. I am used to using Galaxy for analysis but I am trying to go towards programming/command line usage. I am trying to run cufflinks via command line but got the following error:

You are using Cufflinks v2.0.2, which is the most recent release.
Error: cannot open reference GTF file /--GTF for reading



Here is my input text into the command line(could have errors but I am not sure):

cufflinks -G/--GTF /Users/chriscard/Desktop/Galaxy2-\[UCSC_Main_on_Cow__all_mrna_\(genome\)\].gtf /Users/chriscard/Desktop/Galaxy14-\[Tophat_high_fert__accepted_hits\].bam o- /Users/chriscard/Desktop/Command_Line_Output

I plan on using tophat eventually as the first step to command line data analysis but for now I just tried to pull the .bam file from a tophat run off of one of my successful galaxy runs as well as the .gtf file for the reference mRNA that I want cufflinks to use(this is successful via Galaxy). Is there something wrong with my input text into command line? Do I need a different mRNA .gtf file? Sorry if this seems like a very simple task to be having problems with, trying to dive head first into this command line and coding stuff via a couple books I have started reading.

Thank You,
Chris

[Bukowski]

Good on you for leaving Galaxy and going for it on the command line.

The reason you're seeing this error is actually explained in the error, but if you're not used to command line work it won't seem that way.

-G/--GTF

is being interpreted as passing the 'argument' of /--GTF to the 'flag' -G

When you've read the command line you have assumed that you need -G/--GTF because that's what the manual says (no shame in taking it literally) however -G is the same as --GTF and the "/" just means 'use one or the other'.

You may still run into errors by just using one flag, you seem to be also trying to escape all your bracketed characters (preceding ) and ] with \) and I'm not sure why you feel you need to do this. I'd try avoiding having filenames with these kind of characters in. I'm wondering if you're doing this because it's how the command line appears when you're running in Galaxy?

[GenoMax]

Chris,

Unix commands in general will have the following format:

name_of_command [-option 1 -option 2 -option n]

Some of the commands may not need any options at all.

Most of the times if you type the command, with -h or --help option, then you will get a list of all the possible arguments a command will accept, make a note of the default parameters shown for various cufflinks options by running the following.

cufflinks --help

Check out this guide that will show you how to run the Tophat/Cufflinks suite of commands written by the program's author: http://www.nature.com/nprot/journal/....2012.016.html

[ccard28]

I can get cufflinks to run with just this in the command line: cufflinks highferttophat.bam

However when I try and add the reference genome option in which I am typing this into the command line:

new-hostesktop chriscard$ cufflinks [--GTF cowallmrna.gtf] highferttophat.bam


I get this message which I am not sure how to fix

You are using Cufflinks v2.0.2, which is the most recent release.
open: No such file or directory
File [--GTF doesn't appear to be a valid BAM file, trying SAM...
Error: cannot open alignment file [--GTF for reading

This is the same gtf file I had been using successfully as a reference gtf file to use for cufflinks through Galaxy. It seems like it has a problem with the file but I was wondering if maybe my option typing in the command line has errors I am failing to recognize? Even though this is the reference file I am using for galaxy does it need to be a different format through command line? The file type in the manual is gtf which is what this is but I keep getting this error.


Sorry again, I am very new to this. I appreciate all of the help I have gotten and any more anyone can offer.

Thank You,
Chris

[kcchan]

You don't need the square brackets, so it should look something like this:

cufflinks --GTF cowallmrna.gtf highferttophat.bam