Hi, we just had a 16S (V4) miseq-sequencing run per EMP protocol. The run went fine but when we try to process the samples on qiime preprocessing app, the summary sheet (produced by biom summarize-table) shows too low read count e.g. only 1 to 50 reads/sample. On the other hand, the summary produced by split_libraries_fastq.py in the same ppreprocessing run shows much higher reads/ sample e.g. over 50000 seq/ sample. Summarized table and one sample (RN) details are pasted below. Plz note the 94017 sequence in this sample vs. only 30.0 in the summary over it. We've been doing this for long time without ever facing such error; and we used same protocol, barcodes and library prep methods this time too, so don't really know where's the problem. We cannot proceed to qiime visualization without fixing this problem. Could anyone please help in resolving this? Will be highly grateful for help. Many thanks in advance!
Summary produced by biom summarize-table
Num samples: 7 Num observations: 38 Total count: 150 Table density (fraction of non-zero values): 0.195
Min: 4.0 Max: 39.0 Median: 28.000 Mean: 21.429 Std. dev.: 12.960 Sample Metadata Categories: None provided Observation Metadata Categories: taxonomy
B6Old78wk3: 4.0 B6Old78wk2: 8.0 B6Old78wk1: 9.0 B6Young4wk3: 28.0 RN: 30.0 B6Young4wk1: 32.0 B6Young4wk2: 39.0 Summary produced by split_libraries_fastq.py Input file paths Sequence read filepath: /data/input/samples/115162121/RN_S191_L001_R1_001.fastq.gz (md5: 8f40789c606c389722dc770ef4b3939f) Quality filter results Total number of input sequences: 94104 Barcode not in mapping file: 0 Read too short after quality truncation: 61 Count of N characters exceeds limit: 26 Illumina quality digit = 0: 0 Barcode errors exceed max: 0
Result summary (after quality filtering)
Median sequence length: 300.00 RN 94017 Total number seqs written 94017 ---