How to make a metadata mapping file from indexes only

drmusk · January 21, 2019, 4:21am

Hi colleagues
I previously saw few posts on making a metadata or manifest file when you have only sampledID and indexes. Still I have confusion and I am asking it again.

I have 55 Miseq samples as 110 paired end demultiplexed files. I have i7_index, i5_index against each sample ID in an excel file.

I want to make a mapping file for my samples. I am not sure which barcode (eithwe i7 or i5) I should use for samples indexing. May be I have to use both?
Also I am not sure if barcodes or linker primers are still present in the sample sequences. I am attaching barcodes file and 1 fastq file’s head here.
I saw that in the previous posts Mehrbod_Estaki mentioned to make a manifest file and move-on. But it will be difficult for me to proceed with manifest file.
My request here is that if someone can help me to make a mapping file from the available information and then I can follow atacama soil tutorial for analysis.

Lastly. If making a manifest file is the best option here then what will be the (1) command for importing data and (2) which tutorial I should use for subsequent analysis.

Index file

|Sample_ID|i7_index       |i5_index|
|1                |ACCTGGAT|TTCTAGAC|
|2                |ACCTGGAT|CCGATCTT|
|3                |ACCTGGAT|TAAGATCC|
|4                |ACCTGGAT|AGGTCATT|

FASTQ head

@M00779:243:000000000-B8HC8:1:1101:9345:1087 1:N:0:44
CNTACGGGTGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCCATGCCGCGTGTATGAAGAAGGCCTTCGGGTTGTAAAGTACTTTCAGCGGGGAGGAAGGTGTTGAGGTTAATAACCTCAGCAATTGACGTTACCCGCAGAAGAAGCACCGGCTAACTCCGTGCCAGCAGCCGCGGTAATACGGAGGGTGCAAGCATTAATCGGAATTACTGGGCGTAAAGCGCACGCAGGCGGTCTGTCAAGTCGGATGGGAAATCCCCGGGCTCAACCTGGGAACTGCATTCGAA
+
C#[email protected]@FGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGGGGGGFGGGFFGGGGGGGGGGGGGGGEFGFGGGGGGGGFEGGGGGGGGGGGCGGGGGGGGGGDEGGGFGGG,[email protected]<F,CFGGCGGGGGGGGGGGGCFF7FGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGEGEGGGGGGGECEECCFGGFGGFGGGGGGGGGFCCFGGG*):;C7FGFGFDG3=CDECEGGG?:?C9<B<=9BD)<488:DGGGGGC(,;CFGF6A>[email protected])9<<AFCC+;
@M00779:243:000000000-B8HC8:1:1101:19854:1090 1:N:0:44
CNTACGGGAGACAGCAGTGGGGAATATTGCACAATGGGGGAAACCCTGATGCAGCAACGCCGCGTGAGTGATGACGGTCTTCGGATTGTAAAGCTCTGTCTTTGGGGACGATAATGACGGTACCCAAGGAGGAAGACACGGCTAACTACTGGCCAGTAGCCGCGGTCATACGTAGGTGGCAAGCGTTGTCCGGATTTACTGGGCGTAAAGGTAGCGTAGGCGGATTTTTAAGTGGGATGCGCAATACCCGGGCTCACACTGGGTGCTGAGTTCCAAACTGGGACTGGGGAGTGCAGGCGGG
+
C#[email protected]<[email protected],@[email protected]<EFEFDG<EFC<[email protected]++:[email protected]<<[email protected]@[email protected]>[email protected]<++,:,B<FFF++8BD<CF,338,3+,3,[email protected]<F*F1*4,@F,,,>7,782<,;:B4:4=CCBF=:1CBCCFFG81*=*1<92>+0:2*2*9CC*8*3,*/*300A>*95)9*<*:*+0<EE5728C++/0?14*057<9*[email protected]+<<:<968**2***(0C5C<:<C)2*:
@M00779:243:000000000-B8HC8:1:1101:16182:1105 1:N:0:44
CCTACGGGTGGCTGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCCATGCCGCGTGTATGAAGAAGGCCTTCGGGTTGTAAAGTACTTTCAGCGGGGAGGAAGGCGATAAGGTTAATAACCTTGTCGATTGACGTTACCCGCAGAAGAAGCACCGGCTAACTCCGTGCAAGCAGCCGCGGTAATACGGAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAAGCGCACGCAGGCGGTCTGTCAAGTCGGATGTGAAATCCCCGGGCTCAACCTGGGAACTGCATTCGAA
+
[email protected]EGGGG9FFFGFGGGGGGFFGGFFG:DFGGGGGDGGGGGGECGGGCFFGGGGGEEGGGGGGGGG<FFGGGCF9FEEC<FGGEFCGGGDC33*@FDDGGGGGGGGGGGFGGGGGGG*?CGGGGGCEEGGGGGGFBC*BDGEEDCCCCCEE8F7:CEGGGGGGGFGG<E58CFFFG>GFGGFDGBGG8E8*+:A7+09C=93//8:[email protected](3;E902<@CC7;89F<7)<FCCC
@M00779:243:000000000-B8HC8:1:1101:10951:1109 1:N:0:44
CCTATGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCCACGCCGCGTGAGTGAAGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGTGAAGAATAGTGATAAGAGTAACTGCTTATCATGTGACGGTAGCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGAAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGCGCGCGCAGGCGGTCTTTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGGCATGGGAA
+
[email protected]FFEFGGG9FGGGGGFFGGGGGGGGGGGGFGGGGGGGGGGGG<FEEFFF9D9FGDCF9?FGGGFCFGGFDG<FGGFGGFGGGGF+BFGGGGGGFGGGGFGGGGGEFEGGFGGGGGGGGGG>CFGGGGGCFGGGD*,=CFFGGGGGGGG*[email protected]?CG9FCCED5;CCFGFGEGD>G?:EDECEGFE7<[email protected]+:7FGE88>;0;@@G6CEGC(<C))--3*020+
@M00779:243:000000000-B8HC8:1:1101:17684:1109 1:N:0:44
CCTACGGGAGACAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCCATGCCGCGTGTATGAAGAAGGCCTTCGGGTTGTAAAGTACTTTCAGCGGGGAGGAAGGTGTTGAGGTTAATAACCTCAGCAATTGACGTTACCCGCAGAAGAAGCACCGGCTAACTCCGTGCCAGCAGCCGCGGTAATACGGAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAAGCGCACGCCGGCGGTCTGTCACGTCGGATGTGAAATCCCCGGGCTCAACCTGGGAACTGCATTCGAA
+
CCCCCGCGGGGGGGGFGEFGGGGGGGGGFGFGGGGGGGGGGGGGFGFFGGGGGGGGGAEGGGGGFGGGGGGGGEGGGGGGGFFEG<?FGFFFFGGGGGFGGGGGGGGDEGCBFGGGGGGGGGGGGFEFFGGFGGGGGGGGGGGDGG<EFFGGGGGGGGFGGG>EGDGG:[email protected]@FGEEGGGGGGGGG?CGGGGGGGGGGDGFGDDEG3)::5?C*28C:9C+*;5):1:CFF7FFFGGGFGGE0(;CFFG?CCC([email protected]?F?FFF*[email protected]

thermokarst · January 22, 2019, 4:47pm

Let’s take a step back — are your reads multiplexed, or demultiplexed (per-sample?).

drmusk · January 22, 2019, 5:41pm

Dear Dr Dillon

My samples are demultiplexed. As I told that I have 110 fastq.qz files for 55 samples.

thermokarst · January 22, 2019, 5:44pm

Cool! So then, as far as QIIME 2 goes, you don’t need your barcodes in a metadata file at all, since those would only be used for the purposes of demultiplexing (and as you just showed, that isn’t necessary).

Your manifest file (which is different than a metadata file), will reference the files you just shared above, and that is it!

drmusk · January 23, 2019, 8:08am

Dear Dr Dillon
I have made manifest file as it is below.
Now I assume I need to run this command to import sequences

qiime tools import
–type ‘SampleData[PairedEndSequencesWithQuality]’
–input-path manifest_zubi.cvs
–output-path paired-end-demux.qza
–source-format PairedEndFastqManifestPhred64

Wherein “manifest_zubi.cvs” is my manifest file name
source-format may be “PairedEndFastqManifest” but it still doesnt work

AND This command gives error message “Error: no such option: --source-format”

sample-id,absolute-filepath,direction
1,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_1_R1.fastq.gz,forward
1,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_1_R2.fastq.gz,reverse
2,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_2_R1.fastq.gz,forward
2,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_2_R2.fastq.gz,reverse
3,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_3_R1.fastq.gz,forward
3,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_3_R2.fastq.gz,reverse
4,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_4_R1.fastq.gz,forward
4,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_4_R2.fastq.gz,reverse

thermokarst · January 23, 2019, 3:24pm

thermokarst · January 23, 2019, 4:25pm

Great @drmusk!

By the way, I am not a Doctor, myself.

It sounds like you are using docs that don't match your version of QIIME 2 installed - in recent versions of QIIME 2 the parameters in this command have been renamed, in particular, --source-format was renamed to --input-format. If you ever run into a problem like this it helps to run the command with the --help flag to read the latest help text!

thermokarst · January 23, 2019, 4:25pm

drmusk · January 24, 2019, 4:46pm

Dear Colleague
I used below command to import demultiplexed fastq,qz files. but it gave me error message.

$ qiime tools import --type ‘SampleData[PairedEndSequencesWithQuality]’ --input-path Zubi_manifest_4.csv --output-path paired-end-demux.qza --input-format PairedEndFastqManifestPhred33
There was a problem importing Zubi_manifest_4.csv:

Zubi_manifest_4.csv is not a(n) PairedEndFastqManifestPhred33 file:

Found header on line 1 with the following labels: [’“sample-id’, ‘absolute-filepath’, 'direction”’], expected: [‘sample-id’, ‘absolute-filepath’, ‘direction’]

I am attaching my manifest file here.
Can you please help me to build this command?
May be I also need to rename --input-format ?

sample-id,absolute-filepath,direction
zr1577-1,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_1_R1.fastq.gz,forward
zr1577-1,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_1_R2.fastq.gz,reverse
zr1577-2,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_2_R1.fastq.gz,forward
zr1577-2,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_2_R2.fastq.gz,reverse
zr1577-3,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_3_R1.fastq.gz,forward
zr1577-3,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_3_R2.fastq.gz,reverse
zr1577-4,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_4_R1.fastq.gz,forward
zr1577-4,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_4_R2.fastq.gz,reverse
zr1577-5,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_5_R1.fastq.gz,forward
zr1577-5,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_5_R2.fastq.gz,reverse
zr1577-6,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_6_R1.fastq.gz,forward
zr1577-6,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_6_R2.fastq.gz,reverse
zr1577-7,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_7_R1.fastq.gz,forward
zr1577-7,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_7_R2.fastq.gz,reverse
zr1577-8,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_8_R1.fastq.gz,forward
zr1577-8,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_8_R2.fastq.gz,reverse
zr1577-9,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_9_R1.fastq.gz,forward
zr1577-9,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_9_R2.fastq.gz,reverse
zr1577-10,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_10_R1.fastq.gz,forward
zr1577-10,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_10_R2.fastq.gz,reverse
zr1577-11,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_11_R1.fastq.gz,forward
zr1577-11,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_11_R2.fastq.gz,reverse
zr1577-12,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_12_R1.fastq.gz,forward
zr1577-12,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_12_R2.fastq.gz,reverse
zr1577-13,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_13_R1.fastq.gz,forward
zr1577-13,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_13_R2.fastq.gz,reverse
zr1577-14,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_14_R1.fastq.gz,forward
zr1577-14,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_14_R2.fastq.gz,reverse
zr1577-15,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_15_R1.fastq.gz,forward
zr1577-15,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_15_R2.fastq.gz,reverse
zr1577-16,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_16_R1.fastq.gz,forward
zr1577-16,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_16_R2.fastq.gz,reverse
zr1577-17,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_17_R1.fastq.gz,forward
zr1577-17,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_17_R2.fastq.gz,reverse
zr1577-18,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_18_R1.fastq.gz,forward
zr1577-18,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_18_R2.fastq.gz,reverse
zr1577-19,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_19_R1.fastq.gz,forward
zr1577-19,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_19_R2.fastq.gz,reverse
zr1577-20,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_20_R1.fastq.gz,forward
zr1577-20,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_20_R2.fastq.gz,reverse
zr1577-21,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_21_R1.fastq.gz,forward
zr1577-21,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_21_R2.fastq.gz,reverse
zr1577-22,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_22_R1.fastq.gz,forward
zr1577-22,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_22_R2.fastq.gz,reverse
zr1577-23,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_23_R1.fastq.gz,forward
zr1577-23,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_23_R2.fastq.gz,reverse
zr1577-24,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_24_R1.fastq.gz,forward
zr1577-24,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_24_R2.fastq.gz,reverse
zr1577-25,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_25_R1.fastq.gz,forward
zr1577-25,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_25_R2.fastq.gz,reverse
zr1577-26,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_26_R1.fastq.gz,forward
zr1577-26,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_26_R2.fastq.gz,reverse
zr1577-27,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_27_R1.fastq.gz,forward
zr1577-27,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_27_R2.fastq.gz,reverse
zr1577-28,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_28_R1.fastq.gz,forward
zr1577-28,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_28_R2.fastq.gz,reverse
zr1577-29,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_29_R1.fastq.gz,forward
zr1577-29,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_29_R2.fastq.gz,reverse
zr1577-30,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_30_R1.fastq.gz,forward
zr1577-30,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_30_R2.fastq.gz,reverse
zr1577-31,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_31_R1.fastq.gz,forward
zr1577-31,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_31_R2.fastq.gz,reverse
zr1577-32,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_32_R1.fastq.gz,forward
zr1577-32,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_32_R2.fastq.gz,reverse
zr1577-33,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_33_R1.fastq.gz,forward
zr1577-33,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_33_R2.fastq.gz,reverse
zr1577-34,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_34_R1.fastq.gz,forward
zr1577-34,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_34_R2.fastq.gz,reverse
zr1577-35,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_35_R1.fastq.gz,forward
zr1577-35,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_35_R2.fastq.gz,reverse
zr1577-36,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_36_R1.fastq.gz,forward
zr1577-36,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_36_R2.fastq.gz,reverse
zr1577-37,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_37_R1.fastq.gz,forward
zr1577-37,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_37_R2.fastq.gz,reverse
zr1577-38,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_38_R1.fastq.gz,forward
zr1577-38,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_38_R2.fastq.gz,reverse
zr1577-39,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_39_R1.fastq.gz,forward
zr1577-39,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_39_R2.fastq.gz,reverse
zr1577-40,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_40_R1.fastq.gz,forward
zr1577-40,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_40_R2.fastq.gz,reverse
zr1577-41,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_41_R1.fastq.gz,forward
zr1577-41,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_41_R2.fastq.gz,reverse
zr1577-42,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_42_R1.fastq.gz,forward
zr1577-42,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_42_R2.fastq.gz,reverse
zr1577-43,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_43_R1.fastq.gz,forward
zr1577-43,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_43_R2.fastq.gz,reverse
zr1577-44,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_44_R1.fastq.gz,forward
zr1577-44,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_44_R2.fastq.gz,reverse
zr1577-45,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_45_R1.fastq.gz,forward
zr1577-45,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_45_R2.fastq.gz,reverse
zr1577-46,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_46_R1.fastq.gz,forward
zr1577-46,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_46_R2.fastq.gz,reverse
zr1577-47,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_47_R1.fastq.gz,forward
zr1577-47,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_47_R2.fastq.gz,reverse
zr1577-48,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_48_R1.fastq.gz,forward
zr1577-48,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_48_R2.fastq.gz,reverse
zr1577-49,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_49_R1.fastq.gz,forward
zr1577-49,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_49_R2.fastq.gz,reverse
zr1577-50,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_50_R1.fastq.gz,forward
zr1577-50,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_50_R2.fastq.gz,reverse
zr1577-51,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_51_R1.fastq.gz,forward
zr1577-51,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_51_R2.fastq.gz,reverse
zr1577-52,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_52_R1.fastq.gz,forward
zr1577-52,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_52_R2.fastq.gz,reverse
zr1577-53,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_53_R1.fastq.gz,forward
zr1577-53,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_53_R2.fastq.gz,reverse
zr1577-54,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_54_R1.fastq.gz,forward
zr1577-54,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_54_R2.fastq.gz,reverse
zr1577-55,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_55_R1.fastq.gz,forward
zr1577-55,/media/sf_qiime2.data/zubi/Zubair_Bats/zr1577_55_R2.fastq.gz,reverse

thermokarst · January 24, 2019, 7:44pm

This error suggests that your manifest file has quotes (") at the start and end of each line — Microsoft Excel sometimes does this when you save. Maybe you can change the settings when saving? Or, copy and paste the contents into a new file.

drmusk · January 24, 2019, 8:15pm

Thanks for replying
Apparently, it doesn’t show if it has quotes over it. But I will try something.
The other thing. Is below input-format script right? Or I should try both Phred33 and Phred64.

–input-format PairedEndFastqManifestPhred33

drmusk · January 25, 2019, 6:27am

Dear Mathew
Thanks
It worked.
I actually saved my manifest file as txt file and it worked well.
The output .qza file is exactly of the same size as was my compressed fastq.qz

thermokarst · January 25, 2019, 4:08pm

An off-topic reply has been split into a new topic: Questions about QIIME 2 Metadata

Please keep replies on-topic in the future.

system · February 25, 2019, 10:08pm

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