Hi all,
as asked above, do you think would it be possible or would there be any significant reason for the inclusion of singletons in the assembly?
Literally, zero read contigs does not make sense.
I assembled 25M, 90bp fastq reads using trinity assembler and obtained 0.13 Million contigs/transcripts.
I have mapped fastq reads to those transcripts i got by assembling the reads using trinity.
Now when i found the number of reads mapped to each contig/transcript using bowtie2 and samtools, i am confused to see few contigs also had 1 read as well as 0 read.
I also observed that these 1 read, 0 read contig had length of 100bp-500bp length. If the reads are not overlapped into contigs why would the length be >90bp. But, my question is why did bowtie2 showed 0 reads for few of the contig still?
All ideas are welcome and help is appreciated.
Thanks in advance.
+1. Yes. This is a better explanation. Although what I just said is not wrong, it should have a smaller effect. I am retracting my answer. Yours is the correct one.
+1. Yes. This is a better explanation. Although what I just said is not wrong, it should have a smaller effect. I am retracting my answer.
Thank you Jeremy,lh3. Yes i agree that kmers are used in the assembly. But could i please know why i am getting transcripts without any kmer? I used a kmer of 43 in velvet and used inbuilt (.afg) file to get the following results. The numbers below show Contig_ID, Reads( must be kmers from different reads),length of the contig.
93764 0 87 95957 0 88 105528 0 89 509 1 85 2015 1 85 6601 1 85 . . . 897 2 85 2728 2 87 4386 2 85 . .
How is it possible that the length of contig becomes 87 without any overlap with other kmers? Please clarify and help me if i am misunderstanding.
did you actually turn on read tracking?
what was your velvetg command?
Hi Jeremy. I used the following commands to assemble: ./velveth dir 39,65,4 -fastq -short s_read_1.fq s_read_2.fq ./velvetg dir_43 -read_trkg yes -amos_file yes -unused_reads yes
we would have to see the afg file. AFG is a pretty weird format in that scaffolds and contigs are treated similarly. Make sure you review the spec carefully. http://biostar.stackexchange.com/questions/10745/questions-about-the-asm-file-produced-by-the-velvet-program