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  • what are acceptable N50 values for a bacterial genome (assembly using A5)

    I recently paired-end sequenced ~8Mbp bacterial genome using Illumina HiSeq. FastQC quality statistics boxplots are attached both R1 (forward) and R2 (reverse).
    I used A5 to assemble a bacterial genome because of it's automated data cleaning, error correction, contig assembly, scaffolding and quality control. Using A5, one can input the raw reads, which I find a big help, and also that they support the Nextera prep protocol in their quality control.

    My question is, what is an acceptable N50 value? I know that we should aim for high values, which would indicate good assembly. But what is the range of acceptable N50 and what is considered high? Is there a minimum number that we should aim for?

    My A5 summary file says:
    After scaffolding raw1:
    Total number of scaffolds = 5782
    Sum (bp) = 7350401
    Max scaffold size = 54663
    Min scaffold size = 254
    Average scaffold size = 1271
    N50 = 1820

    Any help and feedback is greatly appreciated! Thanks!
    Attached Files

  • #2
    I forgot to add in my previous post that our coverage is ~140x.

    Comment


    • #3
      I don't know the A5 pipeline, but given your coverage and read quality, I think an N50 under 2kb, and a max size at 55 kbp is not too good. I would try SPADES...

      Comment


      • #4
        A case for miseq run?

        Originally posted by CherylM View Post
        I recently paired-end sequenced ~8Mbp bacterial genome using Illumina HiSeq. FastQC quality statistics boxplots are attached both R1 (forward) and R2 (reverse).
        My question is, what is an acceptable N50 value?
        My A5 summary file says:
        After scaffolding raw1:
        Total number of scaffolds = 5782
        Sum (bp) = 7350401
        Max scaffold size = 54663
        Min scaffold size = 254
        Average scaffold size = 1271
        N50 = 1820
        This looks quite a messy assembly to me (even with 100bp reads)...
        Did you check for the KMER/GC content deviation in fastQC?
        Trimmomatic?
        Was it on V2 illumina (2009-2011) chemistry? (I assume it is GC-rich one).
        I would try doing this one on miseq 2x300bp to 90-180X coverage...



        and try other assemblers...

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