No such file or directory at AftrRAD.pl line 513

[quy...@gmail.com]

Dear Mike,
I'm trying our AftrRAD in addition to running STACKS because there are some parts that wasn't working for me in STACKS.  I have demulitplexed my data using STACKS and this is the command I used to run AftrRAD: perl AftrRAD.pl P2-noP2 dplexedData-1
And these are the things that were printed to the terminal window:

Formatting data file QQ174.fq ...(one line of this for each of my 48 samples)
Arguments entered are...
re    TGCAGG
stringLength    15
BarcodePath    Barcodes/
P2    noP2
minDepth    5
minQual    20
dplexedData    1
Help    0
MaxH    90
Phred    33
minParalog    5
numIndels    3
DataPath    Data/
minIden    90


Running AftrRAD...

Recognized 2 data files.

File names are...
 Barcodes_sample_names AllSamples.txt

Use of uninitialized value $TabSeparatedArray[0] in hash element at AftrRAD.pl line 484, <CURRENTBARCODEFILE> line 49.
Use of uninitialized value $Barcode in hash element at AftrRAD.pl line 498, <CURRENTBARCODEFILE> line 49.
Use of uninitialized value $CurrentFileBarcodes[48] in join or string at AftrRAD.pl line 506.
The barcodes you entered for data file 1 (file Barcodes_sample_names) are...
*ALL OF MY BARCODES HERE*

Use of uninitialized value $CurrentFileSampleNames[48] in join or string at AftrRAD.pl line 507.
The names you entered for data file 1 (file Barcodes_sample_names) are...
*SAMPLE NAMES*

No such file or directory at AftrRAD.pl line 513.

I'm not sure what that last line mean and I couldn't really find an answer from previous posts since I don't have any files in my TempFiles folder except a MasterBarcodeFile.txt and the AllSamples file in my Data.

Any help would be greatly appreciated!
Thank you,
Quynh

[Mike Sovic]

Hi Quynh,

It appears you had a file named 'Barcodes_sample_names' in the 'Barcodes' directory at the beginning of the run.  For a demultiplexed run, you'll probably want to start with an empty Barcodes directory.  If you set the dplexedData flag to '1', the program will create the Barcodes file that it needs.  Give this a try, and let me know if you still have a problem.

                   Mike

[quy...@gmail.com]

Hi Mike,
I got rid of the barcode file and things seemed (?) to run smoothly:
"...
Filtering sequence 288000000.
Use of uninitialized value $TotalBarcodeNonMatches in concatenation (.) or string at AftrRAD.pl line 957.
Use of uninitialized value $TotalBarcodeNonMatches in concatenation (.) or string at AftrRAD.pl line 958.
Demultiplexing all samples for datafile AllSamples.txt.
Killed."

Is it supposed to say "killed" at the end?  I tried running Genotype.pl next but there was an error in the Genotypes.txt file and "No such file or directory at Genotype.pl line 246, <GENOTYPERSCRIPT> line 95."

I have the following files in my TempFiles:
AllGoodSeqsWithAssignableBarcodes1.txt
BarcodeCountsToPlot.txt
BarcodeNamesToPlot.txt
Genotypes.txt (empty)
GoodSeqsNoBarcodes.txt
MasterBarcodeFile.txt

Thank you,
Quynh

[Mike Sovic]

Hi Quynh,

No, I don't think you should get the "killed" message.  Never seen it before, but my first guess is it may be a memory-related issue (RAM).  It seems to have crashed at the most memory-intensive point.  Check the post in Dec. about memory usage and see if it looks like this may be an issue.  I promised a updated version in that post that reduces memory requirements, and still haven't made it available, but I will do that very soon (probably in the next few days).

                 Mike