折騰這么多都是白瞎，STAR就有輸出沒有別對上的pair-end reads的功能
參見：How To Filter Mapped Reads With Samtools
I had the same issue but with Paired End Reads, and I solved using samtools and bamToFastq. You can find bamToFastq here: https://code.google.com/p/hydra-sv/
- If you need unmappedR1.fastq (containing both paired and unpaired R1 unmapped reads) and unmappedR2.fastq ( containing both paired and unpaired R2 unmapped reads).
Use samtools -f 4 to extract all unmapped reads :
samtools view -b -f 4 file.bam > file_unmapped.bam bamToFastq -bam file_unmapped.bam -fq1 unmappedR1.fastq -fq2 unmappedR2.fastq
- If you need unmappedpairedR1.fastq (containing only paired R1 unmapped reads) and unmappedpairedR2.fastq ( containing only paired R2 unmapped reads). Meaning you need all paired reads where at least one of them is unmapped.
Use samtools -F 2 to discard only reads mapped in proper pair:
samtools view -b -F 2 file.bam > file_unmapped.bam bamToFastq -bam file_unmapped.bam -fq1 unmappedpairedR1.fastq -fq2 unmappedpairedR2.fastq