I'm recently trying to perform an RNA seq data analysis and in 1st step, I faced a few questions in my mind, which I would like to understand. Please help to understand these questions.
1) In 1st image, raw data from NCBI-SRA have marked 1&2 at the ends of the reads, What is the meaning of this? are those meaning forward and reverse reads?
2) In the second image I was trying to perform trimmomatic with this data set. I chose "paired-end as a collection" but it does not take any input even though my data was there in "fastqsanger.gz" format. Why is that? Should I treat this paired-end data as single-end data while performing Trimmomatic?
3) in the 3rd and 4th images, I collected the same data from ENA where they give two separate files for 1 and 2 marked data in SRA. Then I tried to process them in Trimmomatic by using "Paired-end as individual dataset" and then run it. Trimmomatic gives me 4 files for those, Why is that? which one will be useful for alignment ??
A big thank you in advance :)
I would highly recommend you that before directly jumping into using certain tools for the analysis, please try to understand the basics behind the data, data types & structures, whys and whats of data processing. And for galaxy, it is a very good platform with very good tutorials. Please go through tutorial before asking question which can easily be solved with minimal self inputs.
For NGS sequencers that have pair-end capability, those 1's and 2's refer to which reads they originate from, Read 1 & Read 2 or Forward & Reverse Read s (https://www.illumina.com/science/technology/next-generation-sequencing/plan-experiments/paired-end-vs-single-read.html). That makes it convenient to have it attached at the end to see what kind of reads you are dealing with before processing the reads. In a similar fashion, within the FASTQ format specification, you also specify whether that particular read belongs to Read 1 or Read 2 (see Illumina sequence identifiers: https://en.wikipedia.org/wiki/FASTQ_format). Regarding your "fastqsanger.gz" format data, is this Sanger sequencing related data? These tools are developed for NGS applications. Regarding the output file, check: https://www.biostars.org/p/199938/ and "Trimmomatic output files" on google.
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