Problem with importing data

[Shruti]

Hi,
I started off ewith WGS data. I used sortmeRNA to extract 16S from my WGS. The output file of sortmeRNA is supposed to be my input file for qiime2.
However, qiime2 is not importing my data file saying its neither phred64 nor 33

Here are the details:

1. The command:
   (qiime2-2018.8) [shruti@login-c01 QIIME2]$ qiime tools import --type ‘SampleData[PairedEndSequencesWithQuality]’ --input-path manifest.csv --output-path /scratch/shruti/QIIME2/samples --input-format PairedEndFastqManifestPhred64
   There was a problem importing manifest.csv:

manifest.csv is not a(n) PairedEndFastqManifestPhred64 file

2.My manifest file:
sample-id,absolute-filepath,directon
10-12,/scratch/shruti/QIIME2/10_12_rRNA_R1.fastq,forward
10-12,/scratch/shruti/QIIME2/10_12_rRNA_R2.fastq,reverse
6-8,/scratch/shruti/QIIME2/6_8_rRNA_R1.fastq,forward
6-8,/scratch/shruti/QIIME2/6_8_rRNA_R2.fastq,reverse

Head of my sortmeRNA output fastq files (paired end) which will be my input to qiime2:
(qiime2-2018.8) [shruti@login-c01 QIIME2]$ head 10_12_rRNA_R1.fastq 10_12_rRNA_R2.fastq
==> 10_12_rRNA_R1.fastq <==
@D00172:482:H57FKBCX2:2:1101:1355:2036 1:N:0:GCCAAT
GGGCTAGTACATTCGGCAGGTGAGTTGTTACACAGTCCTTAGCGGATTCCGACTTCCATGGCCACCGTCCTGCTGTCAAGATGTACTAACACCTTTTGTGG
+
DDDCDHHIHIFHHIHIHHHIIIGGGHHIIHHH?FHIFGFHFCHHICHHGHCC<CHHHHCHHFHEEGFFEHHGEH@GCCGGFHHHHEH?HEFEHHIGIHIHI
@D00172:482:H57FKBCX2:2:1101:1423:2039 1:N:0:GCCAAT
CTGCTTACCAAAAATGGCCCACTAGAAACTCTCAATCGCCAAGCGGTCCAATTAAGAGACGGCTTGTTCTTACATATTTAAAGTTTGGAGAATAGGTTAAG
+
DDDC@HHIIIECEHHHFHIGFHIIIHIIHGD<FHIIHIGHHHIIHIIIIIIHH1<<DFF1EHHIIGIIIHHHEHH@GHI?GEHCGH1DCF1DF?HH1GHIE
@D00172:482:H57FKBCX2:2:1101:1478:2042 1:N:0:GCCAAT
CCCCAGCACGACGGAGTTTAACAAGATTACCCGGGCCTTTCGGCCAAGGAAGTACTCGCTGGCTCCGTCAGTGTAACGCGCGTGCGGCCCAGAACATCTAA

==> 10_12_rRNA_R2.fastq <==
@D00172:482:H57FKBCX2:2:1101:1355:2036 2:N:0:GCCAAT
NNNNNNNATTCGAATGTATCAACACTAGTGGGCCATTTTTGGNANNNNGANNNGNNNATNNGGGATGAACCGAACGCGNNGTTAAGGTGCCNGAGTAGNCN
+
#######<<<DGHGHIIICEEHIH@FCHHIFHIIGHHHH?E<#<####<<###<###<<##<<<<DHH@HHECEHHII##<<<DHHHHHFH#<<EECH#1#
@D00172:482:H57FKBCX2:2:1101:1423:2039 2:N:0:GCCAAT
NNNNNNNGACTAATCGAACCATCTAGTAGCTGGTTCCTGCCGAAGNNNCCNNNAGNNTANCAGAAACTCATATCAGATTTATGTGGTAAAGCGAATGATTA
+
#######<<<<EHHGEECEHE?CCGHHFCEEHIHFFIHIFHIIGH###<<###<<##<<#<<DGEG@GE1<CC1CG1DHEFGHFG@GFHHHHHFHIIGHHG
@D00172:482:H57FKBCX2:2:1101:1478:2042 2:N:0:GCCAAT
NNNNTCNGTGATGCCCTTAGATGTTCTGGGCCGCACGCGCGTTACNNTGANNNAGCNAGCGAGTACTTCCTTGGCCGAAAGGCCCGGGTAATCTTGTTAAA

Can you please look into this?
I am just not being able to spot the flaw!

Thank you in Advance.

Shruti Kutmutia
RA
NTU, Singapore.

[Nicholas_Bokulich]

Hi @Shruti,
Invisible characters can cause issues with the manifest file — see these posts and let us know if you are still having trouble (with details on your troubleshooting) after giving those a try. Thanks!

[Shruti]

Yes
Well I did strip off the BOM characters using vim :set nobomb and many other ways like tail and sed but it does not seem to work.

Here is my manifest file

manifest_1.csv (257 Bytes)

[ebolyen (Evan Bolyen)]

Hi @Shruti,

It looks like its a (very hard to see) typo in your header.
Change directon to direction and I think it will work

Good luck!