Hi,
I've received my microbiome sequencing results from uBiome and I am trying to analyze the raw reads using QIIME 1.9.1. The files are as follows:
ssr_100__R1__L001.fastq.gz,
ssr_100__R1__L002.fastq.gz,
ssr_100__R1__L003.fastq.gz,
ssr_100__R1__L004.fastq.gz,
ssr_100__R2__L001.fastq.gz,
ssr_100__R2__L002.fastq.gz,
ssr_100__R2__L003.fastq.gz,
ssr_100__R2__L004.fastq.gz.
I have tried merging the R1 files and R2 files using cat and then use the join_paired_ends.py script, but only a fraction of reads get joined. Could anyone advise me on propper sequence of steps to analyze this data to perform OTU picking? Perhaps someone has a script to run the automated pipeline for the analysis?
Thanks,
People usually "merge" paired end using tools like FLASH.
qiime provides detailed documents for otu-picking and down-stream analyis. e.g., http://nbviewer.jupyter.org/github/biocore/qiime/blob/1.9.1/examples/ipynb/illumina_overview_tutorial.ipynb
the tutorial does not deal with multi lane, paired end demultiplexed samples. I am unsure in the multiple_split_libraries_fastq.py script performs well on this data.
What is the platform with which the reads were generated. You do not have to cat the read 1 and read 2 since according to the join_paired_ends.py, you have to do the following:
The script merges the paired end reads together. I think this way, you will see merged reads together. Run the qiime pipeline and see if you get good OTU's. If the results are not good, different trouble shooting steps can be done.
I have tried using the script yet most of the reads are left unjoined. Any ideas on how to overcome this?
Have you tried other merge options,
This post seems to have many such options,
A link on Biostars
all join*.py scripts produce the same results. Only the multiple_split_libraries_fastq.py produces a single .fna file, but it is a demultiplexing script so I am not sure how it deals with paired end and multi-lane .fastq files from a single sample.