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We are presently working on the sequencing and assembly of a draft genome of a ~1 Gb invertebrate. The preliminary dataset is a HiSeq single lane of PE100, with a 300 bp insert size. Larger insert libraries will follow (first a 5 Kb mate pair library), but we are attempting to assemble the data on hand.
We are performing this initial assembly using SOAPdenovo2. Our usage is as follows:
This was used in conjunction with the following configuration file:
We've noticed one oddity on more than one scaffold we've manually inspected. I expect contigs to represent contiguous sequences, and the contigs file output by SOAPdenovo2 contains gapless sequences, as expected. The scaffolds file contains, as expected, some scaffolds containing stretches of Ns. However, the oddity is that when I examine the contigs components of a given scaffold, the scaffold has gaps where there are none indicated in the contig. In other words, the scaffolding process seems to introduce gaps within contigs, or the contigs have gaps that are not indicated.
Here is one example, take this scaffold:
Its components are as follows:
However, if you manually inspect and place the contigs on the scaffold, you will notice that scaffold indicates Ns / gaps WITHIN two of the three contigs:
I would greatly appreciate any insights on what is happening here. Thanks!
We are performing this initial assembly using SOAPdenovo2. Our usage is as follows:
Code:
nohup SOAPdenovo-63mer all -s config_swiss -K 55 -R -o soap55 1>ass55.log 2>ass55.err &
Code:
max_rd_len=100 [LIB] avg_ins=300 reverse_seq=0 asm_flags=3 rd_len_cutoff=100 rank=1 pair_num_cutoff=3 map_len=32 q1=~/gen/swiss_1_paired.fq q2=~/gen/swiss_2_paired.fq q=~/gen/swiss_1_unpaired.fq q=~/gen/swiss_2_unpaired.fq
We've noticed one oddity on more than one scaffold we've manually inspected. I expect contigs to represent contiguous sequences, and the contigs file output by SOAPdenovo2 contains gapless sequences, as expected. The scaffolds file contains, as expected, some scaffolds containing stretches of Ns. However, the oddity is that when I examine the contigs components of a given scaffold, the scaffold has gaps where there are none indicated in the contig. In other words, the scaffolding process seems to introduce gaps within contigs, or the contigs have gaps that are not indicated.
Here is one example, take this scaffold:
Code:
>scaffold1 10.9 AATAAATATGCATGCCATAAAAAATGCAAGTTACAATACAAATTTTTTAAATTTATAACTAGTTTAAACTTACAATGTGACACCACAGCCATCTATGTAA ATAATAAATTCTAAATTAATACAAACACATCAGATATAAGATTACTTGTAACCTAACTGAATAAATATGCACGCCCAGGGCCGTACCGACGGAGGGGGGT GGGGGTAGACACCCCCCAGATATGCAAAACTCTGTTGGTTTTTGACGAAAAACTCGAAAATTATAACTTCTGTCGGGTAAAACTGAAATTTGGCTTCTTA AAAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTCNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNACCCCCCCCCCACGTCTTTGTCGTTCGGTACGGCCCTGGTT ATAAATCGTTTACCAANCCCCCCCCCCCACGTCTTTGTCGTTTGGTACGGCTCTGTGCTCGCCGAAAGAAATGCAAGTTACAATATAAATTTTAACCAAT TTAAACTTACAGTGACATACATGCCAAATGCCAAGTGTCTAAAATAAAAAATTTTAAAATACCATAAACAATCTATATATAAACATAGTTTTAACACAAC TAATAATTATGAACGTCAAAAGAAATGCACGCTACAATATAATTTTTTTAGTATTACAACTAATTTAAACTTAACAGTATATTAACCACTTGCCACATGT AATAAAATTATGTTATAAATTTAGTTAAATAGAACTAGTTAAAAACTTTATAAAAGGCTTGTTAATAATCAAAATTATAATTTTATTGGGTTGCAACTAA
Code:
>scaffold1 21162015 0 + 356 16946028 459 + 57 21633622 517 + 452 >>21162015 length 356 cvg_17.6_tip_0 AATAAATATGCATGCCATAAAAAATGCAAGTTACAATACAAATTTTTTAAATTTATAACTAGTTTAAACTTACAATGTGACACCACAGCCATCTATGTAA ATAATAAATTCTAAATTAATACAAACACATCAGATATAAGATTACTTGTAACCTAACTGAATAAATATGCACGCCCAGGGCCGTACCGACGGAGGGGGGT GGGGGTAGACACCCCCCAGATATGCAAAACTCTGTTGGTTTTTGACGAAAAACTCGAAAATTATAACTTCTGTCGGGTAAAACTGAAATTTGGCTTCTTA AAAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTC >16946028 length 112 cvg_1.9_tip_0 AAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTCACCCCCCCCCCACGTCTTTGTCGTTCGGTACGGCCCTGGTTATAA ATCGTTTACCAA >21633622 length 507 cvg_8.0_tip_0 AAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTCCCCCCCCCCCCACGTCTTTGTCGTTTGGTACGGCTCTGTGCTCGC CGAAAGAAATGCAAGTTACAATATAAATTTTAACCAATTTAAACTTACAGTGACATACATGCCAAATGCCAAGTGTCTAAAATAAAAAATTTTAAAATAC CATAAACAATCTATATATAAACATAGTTTTAACACAACTAATAATTATGAACGTCAAAAGAAATGCACGCTACAATATAATTTTTTTAGTATTACAACTA ATTTAAACTTAACAGTATATTAACCACTTGCCACATGTAATAAAATTATGTTATAAATTTAGTTAAATAGAACTAGTTAAAAACTTTATAAAAGGCTTGT TAATAATCAAAATTATAATTTTATTGGGTTGCAACTAAGTTTATATTTGCCATACACGCTGGTAAAATTGAGTTAAAAATCTCCCACATCATAGTCCAAA ACAACGT
Code:
>16946028 length 112 cvg_1.9_tip_0 AAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTC [COLOR="Red"]GAP[/COLOR] ACCCCCCCCCCACGTCTTTGTCGTTCGGTACGGCCCTGGTTATAA ATCGTTTACCAA >21633622 length 507 cvg_8.0_tip_0 AAAAAAAAAAAAAAATTATAAGATCTTGTCGGTAAATTTAGCTTTTTACATTTTC [COLOR="Red"]GAP[/COLOR] CCCCCCCCCCCACGTCTTTGTCGTTTGGTACGGCTCTGTGCTCGC CGAAAGAAATGCAAGTTACAATATAAATTTTAACCAATTTAAACTTACAGTGACATACATGCCAAATGCCAAGTGTCTAAAATAAAAAATTTTAAAATAC CATAAACAATCTATATATAAACATAGTTTTAACACAACTAATAATTATGAACGTCAAAAGAAATGCACGCTACAATATAATTTTTTTAGTATTACAACTA ATTTAAACTTAACAGTATATTAACCACTTGCCACATGTAATAAAATTATGTTATAAATTTAGTTAAATAGAACTAGTTAAAAACTTTATAAAAGGCTTGT TAATAATCAAAATTATAATTTTATTGGGTTGCAACTAAGTTTATATTTGCCATACACGCTGGTAAAATTGAGTTAAAAATCTCCCACATCATAGTCCAAA ACAACGT