Open christinalrichards opened 5 years ago
Hi,
I suggest you follow this suggestion: https://github.com/BSSeeker/BSseeker2#qa11 by splitting the large input file into pieces, and then map them independently. The performance will be largely improved.
Best, Weilong
Thank you for your response! Do you think it did not finish or are you concerned about the low mapping efficiency? I should have clarified that this is mapping the wild relative of tomato Lycopersicon pimpinellifolium to the domesticated tomato genome Solanum lycopersicum divergence: Estimated Time: 0.6751 MYA (according to http://www.timetree.org/). Perhaps I should try running one fo the samples of S. lycopersicum.
Also how can I use this code for PE reads "split -l 4000000 input.fq"
For low mapping efficiency, please check to allow more mismatch, or to allow gap alignment, such as using bowtie2-end-to-end alignment mode or bowtie2-local alignment mode.
Split the large file will speed up the total time cost, and let me know whether the command works as early as possible.
Best, Weilong
zcat R1.fastq.gz | split -l 4000000 - R1 zcat R2.fastq.gz | split -l 4000000 - R2
Hi! When I split the reads for R1 I get 24 files: -rw-r----- 1 clr pi_clr 238489543 Sep 6 16:27 xaa -rw-r----- 1 clr pi_clr 238494383 Sep 6 16:27 xab -rw-r----- 1 clr pi_clr 238505358 Sep 6 16:27 xac -rw-r----- 1 clr pi_clr 238516820 Sep 6 16:27 xad -rw-r----- 1 clr pi_clr 238489803 Sep 6 16:27 xae -rw-r----- 1 clr pi_clr 238478038 Sep 6 16:27 xaf -rw-r----- 1 clr pi_clr 238492368 Sep 6 16:27 xag -rw-r----- 1 clr pi_clr 238489493 Sep 6 16:27 xah -rw-r----- 1 clr pi_clr 238484043 Sep 6 16:27 xai -rw-r----- 1 clr pi_clr 238487565 Sep 6 16:27 xaj -rw-r----- 1 clr pi_clr 238483930 Sep 6 16:27 xak -rw-r----- 1 clr pi_clr 238487671 Sep 6 16:27 xal -rw-r----- 1 clr pi_clr 238483939 Sep 6 16:27 xam -rw-r----- 1 clr pi_clr 238487869 Sep 6 16:27 xan -rw-r----- 1 clr pi_clr 238513293 Sep 6 16:27 xao -rw-r----- 1 clr pi_clr 238500051 Sep 6 16:27 xap -rw-r----- 1 clr pi_clr 238513998 Sep 6 16:27 xaq -rw-r----- 1 clr pi_clr 238504783 Sep 6 16:27 xar -rw-r----- 1 clr pi_clr 238494160 Sep 6 16:27 xas -rw-r----- 1 clr pi_clr 238486544 Sep 6 16:27 xat -rw-r----- 1 clr pi_clr 238454100 Sep 6 16:27 xau -rw-r----- 1 clr pi_clr 238507996 Sep 6 16:27 xav -rw-r----- 1 clr pi_clr 238476629 Sep 6 16:27 xaw -rw-r----- 1 clr pi_clr 36872879 Sep 6 16:27 xax
and then if I stay in the same directory and split R2 it overwrites those 24 files with theses 24 files: -rw-r----- 1 clr pi_clr 238489543 Sep 6 16:32 xaa -rw-r----- 1 clr pi_clr 238494383 Sep 6 16:32 xab -rw-r----- 1 clr pi_clr 238505358 Sep 6 16:32 xac -rw-r----- 1 clr pi_clr 238516820 Sep 6 16:32 xad -rw-r----- 1 clr pi_clr 238489803 Sep 6 16:32 xae -rw-r----- 1 clr pi_clr 238478038 Sep 6 16:32 xaf -rw-r----- 1 clr pi_clr 238492368 Sep 6 16:32 xag -rw-r----- 1 clr pi_clr 238489493 Sep 6 16:32 xah -rw-r----- 1 clr pi_clr 238484043 Sep 6 16:32 xai -rw-r----- 1 clr pi_clr 238487565 Sep 6 16:32 xaj -rw-r----- 1 clr pi_clr 238483930 Sep 6 16:32 xak -rw-r----- 1 clr pi_clr 238487671 Sep 6 16:32 xal -rw-r----- 1 clr pi_clr 238483939 Sep 6 16:33 xam -rw-r----- 1 clr pi_clr 238487869 Sep 6 16:33 xan -rw-r----- 1 clr pi_clr 238513293 Sep 6 16:33 xao -rw-r----- 1 clr pi_clr 238500051 Sep 6 16:33 xap -rw-r----- 1 clr pi_clr 238513998 Sep 6 16:33 xaq -rw-r----- 1 clr pi_clr 238504783 Sep 6 16:33 xar -rw-r----- 1 clr pi_clr 238494160 Sep 6 16:33 xas -rw-r----- 1 clr pi_clr 238486544 Sep 6 16:33 xat -rw-r----- 1 clr pi_clr 238454100 Sep 6 16:33 xau -rw-r----- 1 clr pi_clr 238507996 Sep 6 16:33 xav -rw-r----- 1 clr pi_clr 238476629 Sep 6 16:33 xaw -rw-r----- 1 clr pi_clr 36872879 Sep 6 16:33 xax
Should I somehow rename each of the pairs of files (from R1 and R2) eg: -rw-r----- 1 clr pi_clr 238489543 Sep 6 16:27 xaa -rw-r----- 1 clr pi_clr 238489543 Sep 6 16:32 xaa
to then use this series of commands:
bs_seeker2-align.py -1 FN1 -2 FN2 -g genome.fa -o PE.bam -u unmapped
bs_seeker2-align.py -i unmapped_1.fa -g genome.fa -o unmapped_1.bam
Antisense.py -i unmapped_2.fa -o unmapped_2_antisense.fa
bs_seeker2-align.py -i unmapped_2_antisense.fa -g mm9_phage.fa -o unmapped_2.bam
samtools merge merge.bam PE.bam unmapped_[12].bam
To the following command to avoid overwriting the files.
zcat Mate1.fq.gz | split -l 4000000 - R1_ &
zcat Mate2.fq.gz | split -l 4000000 - R2_ &
Best, Weilong
Hi Dr. Guo,
I have made quite some progress through step 1 and step 2, but I get a FAILED, ExitCode 1 after 12h53m of the mapping of PE reads. I used the attached shell file and it looks like it runs through this much :
Output the unmapped reads in PE mode:
python bs_seeker2-align.py -1 10_P_1.fq -2 10_P_2.fq -g AEKE03.fasta --aligner=bowtie2 -o 10_P.bam -u unmapped
Map the unmapped reads in mate 1:
python bs_seeker2-align.py -i unmapped_1.fa -g AEKE03.fasta -o unmapped_1.bam
Convert the unmapped reads in mate 2 to their reverse complementaries:
python Antisense.py -i unmapped_2.fa -o unmapped_2_antisense.fa
Map the unmapped reads in mate 2:
python bs_seeker2-align.py -i unmapped_2_antisense.fa -g mm9_phage.fa -o unmapped_2.bam
But did it finish? It says END in the log file (also attached).
BSSeeker2_Tomato.sh.docx
10_P.bam.bs_seeker2.log