Hi all,
I’m relatively new to working with sequence data generated from Illumina’s MiSeq. I have previously worked with the QIIME pipeline and wanted to try mothur.
I made a 3 column mapping file with the first column being the sample name, the second column being the forward fastq file name, and the third column being the reverse fastq file name.
I was working through the MiSeq SOP with mothur trying to join my paired end reads but am getting this error:
mothur > make.contigs(file=/Users/laurenoconnell/Desktop/Thesis_Data/mothur/stability.files^T)
Using 1 processors. Reading fastq data... [WARNING]: can't find PE12_S1_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE12_S1_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE15_S2_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE15_S2_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE17_S3_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE17_S3_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE12_S4_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE18_S4_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE20_S5_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE20_S5_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE21_S6_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE21_S6_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE23_S7_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE23_S7_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE26_S8_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE26_S8_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE27_S9_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE27_S9_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE92_S10_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE29_S10_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE30_S11_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE30_S11_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE32_S12_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE32_S12_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE33_S13_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE33_S13_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE35_S14_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE35_S14_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE36_S15_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE36_S15_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE38_S16_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE38_S16_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE39_S17_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE39_S17_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE41_S18_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE41_S18_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE42_S19_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE42_S19_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE44_S20_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE44_S20_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE45_S21_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE45_S21_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE47_S22_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE47_S22_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE48_S23_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE48_S23_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE50_S24_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE50_S24_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE51_S25_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE51_S25_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE52_S26_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE52_S26_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE55_S27_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE55_S27_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE56_S28_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE56_S28_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE61_S29_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE61_S29_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE62_S30_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE62_S30_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE64_S31_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE64_S31_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE66_S32_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE66_S32_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE67_S33_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE67_S33_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE68_S34_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE68_S34_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE71_S35_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE71_S35_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE73_S36_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE73_S36_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE74_S37_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE74_S37_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE79_S38_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE79_S38_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE82_S39_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE82_S39_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE83_S40_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE83_S40_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE85_S41_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE85_S41_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE86_S42_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE86_S42_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE87_S43_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE87_S43_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE89_S44_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE89_S44_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE90_S45_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE90_S45_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE92_S46_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE92_S46_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE93_S47_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE93_S47_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE95_S48_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE95_S48_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE96_S49_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE96_S49_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE98_S50_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE98_S50_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE99_S51_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE99_S51_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE101_S52_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE101_S52_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE102_S53_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE102_S53_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE104_S54_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE104_S54_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE105_S55_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE105_S55_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE107_S56_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE107_S56_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE108_S57_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE108_S57_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE110_S58_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE110_S58_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE111_S59_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE111_S59_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE113_S60_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE113_S60_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE114_S61_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE114_S61_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE119_S62_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE119_S62_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE120_S63_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE120_S63_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE122_S64_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE122_S64_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE123_S65_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE123_S65_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE125_S66_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE125_S66_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE126_S67_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE126_S67_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE128_S68_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE128_S68_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE129_S69_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE129_S69_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE131_S70_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE131_S70_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE132_S71_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE132_S71_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE134_S72_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE134_S72_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE136_S73_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE136_S73_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE138_S74_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE138_S74_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE140_S75_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE140_S75_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE141_S76_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE141_S76_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE143_S77_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE143_S77_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE144_S78_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE144_S78_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE146_S79_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE146_S79_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE147_S80_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE147_S80_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE149_S81_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE149_S81_L001_R2_001.fastq, ignoring pair. [WARNING]: can't find PE150_S82_L001_R1_001.fastq, ignoring pair. [WARNING]: can't find PE150_S82_L001_R2_001.fastq, ignoring pair. Done. It took 0 secs to process 0 sequences.
The data that I am using is already demultiplexed and the adapters/barcodes have been trimmed off.
I was able to generate the fasta files doing each sample individually, but I wasn’t able to get them to work in a batch so that the fasta file was only 1 large combined file of all my samples, rather than individual files per sample.
If anyone has any suggestions on what I might be doing wrong and how to fix it I would greatly appreciate it!
Thanks,
Lauren