Hello Biostar community,
I'm currently working on aligning paired-end FASTQ files from a mig-SEQ experiment to a reference genome. After filtering with fastp and removing chloroplast DNA using Bowtie2 with a chloroplast genome as reference, I have forward and reverse reads that don't match in length exactly.
I encountered an issue when using the following Bowtie2 command:
bowtie2 -x <index_prefix> -1 <forward_fastq> -2 <reverse_fastq> -S <output_sam>
The process terminated with an error message: "Error, fewer reads in file specified with -1 than in file specified with -2."
Thus, it seems as if the best course of action is to combine the the forward and reverse reads first, and then just align the combined fastq file to the index. My question is: what is the best way to combine these reads?
I've noticed that some researchers concatenate the forward and reverse reads before alignment. However, this approach seems somewhat crude. If concatenation is indeed a valid step, should I consider taking the reverse complement of the reverse read before concatenating? This way, all reads would be in the same direction, which seems to be an expectation of Bowtie2.
As I'm relatively new to bioinformatics, I apologize if my question is not well-informed or lacks clarity. Any guidance or suggestions would be greatly appreciated.
Thank you in advance for your help.
Best regards,
Alexander
Thank you so much! This is a lot of help. I will try this.