Hi I am trying to make a -bedpe file using bedtools bamtobed and am running into a problem. I get many of the following error messages after this command.
bedtools bamtobed -bedpe -i male_1_sorted_rmdup.bam > male_1_sorted_rmdup.bed
*****WARNING: Query NB551387:262:H3FMVBGXC:3:21408:20039:5256 is marked as paired, but its mate does not occur next to it in your BAM file. Skipping.
If I run samtools flagstat on the bam file input it appears that the mates are properly paired. These are sorted bam files from which I have removed the duplicates Picard. They are otherwise not processed, made with Bowtie2.
samtools flagstat male_1_sorted_rmdup.bam 21276014 + 0 in total (QC-passed reads + QC-failed reads) 0 + 0 duplicates 21276014 + 0 mapped (100.00%:-nan%) 21276014 + 0 paired in sequencing 10638007 + 0 read1 10638007 + 0 read2 21276014 + 0 properly paired (100.00%:-nan%) 21276014 + 0 with itself and mate mapped 0 + 0 singletons (0.00%:-nan%) 0 + 0 with mate mapped to a different chr 0 + 0 with mate mapped to a different chr (mapQ>=5)
Wondering why this may be happening? Thanks!