# make.contigs group file problem **URL:** \ **Category:** Commands in mothur\ **Created:** [February 29, 2016, 5:00pm UTC](https://forum.mothur.org/t/make-contigs-group-file-problem/2733 "2016-02-29T17:00:20Z")\ **Posts on this page:** 3\ **Page:** 1
**Author:** ![juliak](https://avatars.discourse-cdn.com/v4/letter/j/6bbea6/32.png) [@juliak](https://forum.mothur.org/u/juliak)\ **Post date:** [February 29, 2016, 5:00pm UTC](https://forum.mothur.org/t/make-contigs-group-file-problem/2733/1 "2016-02-29T17:00:20Z")
Hello, I am having trouble with my group file right at the beginning of the MiSeq SOP. I am working with Illumina MiSeq V4 reads and I am using mothur v.1.36.1 on windows. I made contigs using forward and reverse fastq files and when I attempted to screen sequences I got the following error message over and over again and about a third of my sequences are thrown out: Your groupfile does not include the sequence please correct. I then took a closer look at the group file (.contigs.groups) and it seems that some sequence names get cut off and as a result there is a shift and the following sequences won’t be recognized (at least that is what I am assuming) here is my code: mothur \> make.contigs(file=test.files, processors=3) Group count: V4\_29 41611 V4\_30 205103 V4\_31 203345 Total of all groups is 450059 mothur \> summary.seqs(fasta=test.trim.contigs.fasta, processors=3) Using 3 processors. Start End NBases Ambigs Polymer NumSeqs Minimum: 1 296 296 0 3 1 2.5%-tile: 1 307 307 0 4 11252 25%-tile: 1 309 309 0 4 112515 Median: 1 310 310 0 4 225030 75%-tile: 1 310 310 1 5 337545 97.5%-tile: 1 317 317 17 7 438808 Maximum: 1 602 602 68 296 450059 Mean: 1 313.655 313.655 2.04722 4.53988 # of Seqs: 450059 mothur \> screen.seqs(fasta=test.trim.contigs.fasta, group=test.contigs.groups, summary=test.trim.contigs.summary, maxambig=0, minlength=307, maxlength=317) Using 3 processors. Your groupfile does not include the sequence M02973\_22\_000000000-AK66Y\_1\_2112\_24715\_6481 please correct. Your groupfile does not include the sequence M02973\_22\_000000000-AK66Y\_1\_2112\_25076\_17339 please correct. Your groupfile does not include the sequence M02973\_22\_000000000-AK66Y\_1\_2112\_25077\_13464 please correct. Your groupfile does not include the sequence M02973\_22\_000000000-AK66Y\_1\_2112\_25083\_10665 please correct. and so on… mothur \> summary.seqs(fasta=test.trim.contigs.good.fasta, processors=3) Using 3 processors. Start End NBases Ambigs Polymer NumSeqs Minimum: 1 307 307 0 3 1 2.5%-tile: 1 308 308 0 4 7301 25%-tile: 1 309 309 0 4 73008 Median: 1 310 310 0 4 146015 75%-tile: 1 310 310 0 5 219022 97.5%-tile: 1 312 312 0 6 284729 Maximum: 1 317 317 0 12 292029 Mean: 1 309.921 309.921 0 4.47276 # of Seqs: 292029 I appreaciate any hints you can give me! Thanks, Julia ---
**Author:** ![westcott](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/westcott/32/18_2.png) [@westcott](https://forum.mothur.org/u/westcott)\ **Post date:** [March 8, 2016, 1:37pm UTC](https://forum.mothur.org/t/make-contigs-group-file-problem/2733/2 "2016-03-08T13:37:39Z")
Is your group file blank? Have you seen this post, [make.contigs output - XXX.contigs.group blank](https://mothur.ltcmp.net/t/make-contigs-output-xxx-contigs-group-blank/2620/1) ---
**Author:** ![juliak](https://avatars.discourse-cdn.com/v4/letter/j/6bbea6/32.png) [@juliak](https://forum.mothur.org/u/juliak)\ **Post date:** [March 15, 2016, 3:16pm UTC](https://forum.mothur.org/t/make-contigs-group-file-problem/2733/3 "2016-03-15T15:16:36Z")
Hello, thanks for your reply. I am using fastq files and my group file was not blank, so that wasn’t the problem. But I am happy to say that since my last post, I was able to get it working. I am not exactly sure what the actual problem was, but I think my path was maybe too long. So after copying all my mothur files directly on the C: drive (C:\mothur) it seems to be working fine now.