make.contigs issue

Hi,

I am on the new side and am having some trouble with the make.contigs command.

make.contigs(ffastq=Undetermined_S0_L001_R1_001.fastq, rfastq=Undetermined_S0_L001_R2_001.fastq, findex=Undetermined_S0_L001_I1_001.fastq, oligos=oligos.txt):

The files get made alright but when I run summary.seqs it doesn’t look good.

Start End NBases Ambigs Polymer NumSeqs
Minimum: 1 250 250 0 3 1
2.5%-tile: 1 366 366 0 4 167227
25%-tile: 1 370 370 0 4 1672262
Median: 1 373 373 1 5 3344524
75%-tile: 1 374 374 3 5 5016786
97.5%-tile: 1 378 378 17 6 6521821
Maximum: 1 502 502 50 240 6689047
Mean: 1 373.703 373.703 2.61363 4.74528

of Seqs: 6689047

I have also tried splitting this step up:
make.contigs(ffastq=Undetermined_S0_L001_R1_001.fastq, rfastq=Undetermined_S0_L001_R2_001.fastq)
fastq.info(fastq=Undetermined_S0_L001_I1_001.fastq)

but the result is the same.

We did 251 x 251bp runs using 515f-926r primers. In the past we used different primers so I am unsure if that is the issue or if I need more information in the oligo file:

barcode CAACACATGCTG none P13097
barcode CATACCGTGAGT none P13015
barcode GTCCATGGTTCG none P13289
barcode ACCATTACCATT none P13067

Needless to say, if I try filter out anything with a maxambig>0 or that is longer than 275bp, everything winds up in the scrap folder.

Thanks for your help!

the 275 bp length filter is specific to v4 (515f, 806r) your product is another 115bp longer

Ahhh. That makes sense. Thanks for pointing that out.