Hi Kory, The problem with this FASTQ block is that the sequence and quality score identifier lines do not match ('SNPSTER6_0679:1:1:1083:939#0/1 run=100908_SNPSTER6_0679_70929AAXX' vs 'SNPSTER6_0679:1:1:1083:939#0/1'), where the identifier for the sequence line has additional text not found on the identifier for the quality score line, which is not valid for the FASTQ format. Alternatively the quality score identifier line could be only a '+', without the sequence identifier. The quality score lines appear to be either illumina or solexa, but it is best to check with the source of the data to be sure: Input ASCII range: 'B'(66) - '_'(95) Input decimal range: 2 - 31 You'll need to upload valid FASTQ files inorder to work with them in Galaxy. Correct examples of your provided read are: @SNPSTER6_0679:1:1:1083:939#0/1 NATTTATGGATAGTTGGGTAGTAGGTGTAAATGTATGTGGTAAAAGGCCTAGGAGATTTGTTGATCCAATAAATATGATTAGGGAAACAA +SNPSTER6_0679:1:1:1083:939#0/1 BIQQIQQQTP[[[[[VVVVQPPPPPTWWWW[[YYTTTOVV____TWVXRWPTQPQWWWWWTOOVV___V_TROOWTWTWTQWQWTTRWRO or @SNPSTER6_0679:1:1:1083:939#0/1 run=100908_SNPSTER6_0679_70929AAXX NATTTATGGATAGTTGGGTAGTAGGTGTAAATGTATGTGGTAAAAGGCCTAGGAGATTTGTTGATCCAATAAATATGATTAGGGAAACAA + BIQQIQQQTP[[[[[VVVVQPPPPPTWWWW[[YYTTTOVV____TWVXRWPTQPQWWWWWTOOVV___V_TROOWTWTWTQWQWTTRWRO or @SNPSTER6_0679:1:1:1083:939#0/1 run=100908_SNPSTER6_0679_70929AAXX NATTTATGGATAGTTGGGTAGTAGGTGTAAATGTATGTGGTAAAAGGCCTAGGAGATTTGTTGATCCAATAAATATGATTAGGGAAACAA +SNPSTER6_0679:1:1:1083:939#0/1 run=100908_SNPSTER6_0679_70929AAXX BIQQIQQQTP[[[[[VVVVQPPPPPTWWWW[[YYTTTOVV____TWVXRWPTQPQWWWWWTOOVV___V_TROOWTWTWTQWQWTTRWRO Please let us know if we can be of further assistance. Thanks for using Galaxy, Dan On Feb 2, 2011, at 2:01 PM, Johnson, Kory (NIH/NINDS) [C] wrote:

Hello,

My account login is: johnsonko@ninds.nih.gov

I am a first time Galaxy user.

I have uploaded my sequences as format “fastq” into Galaxy and would like to next use “Groomer” to output Sanger fastq format so to go on with exploring quality via box plot, deciding on a trim length (if any), and map to genome using bwa or bowtie.

However, I am running into a problem using “Groomer”.

I do not know what format my sequences are per setting the required input parameter.

An example of my sequences is as follows:

@SNPSTER6_0679:1:1:1083:939#0/1 run=100908_SNPSTER6_0679_70929AAXX NATTTATGGATAGTTGGGTAGTAGGTGTAAATGTATGTGGTAAAAGGCCTAGGAGATTTGTTGATCCAATAAATATGATTAGGGAAACAA +SNPSTER6_0679:1:1:1083:939#0/1 BIQQIQQQTP[[[[[VVVVQPPPPPTWWWW[[YYTTTOVV____TWVXRWPTQPQWWWWWTOOVV___V_TROOWTWTWTQWQWTTRWRO

… how to tell if you have: “Sanger”, “Solexa”, “Illumina 1.3+”, etc.

I have tried to submit to “Groomer” different times using these options one at a time and none return with results.

Need help please.

Also, what is the expected time for “Groomer” to return results for a file containing 2.7 million reads.

Thank you … best,

Kory

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Kory R. Johnson, MS, PhD Sr. Bioinformatics Scientist

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