Chris:LabNotes/Microbiome/2011/2011-5-2

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Revision as of 21:29, 2 May 2011 by >Cjwei
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Slide Shortening Assembly Test

Background

  • Sam tried to increase SAG contig length by using recruited sequences from metagenomic contigs (http://genome-tech.ucsd.edu/LabNotes/index.php/Sam:LabNotes/Microbiome-new/2011-4-19)
    • But results showed some decreased contig lengths after recruitment (ID2, ID9, ID11)
    • Dr. Zhang suggested that the decreased contig lengths may be fixed by slide shortening of contigs
  • On 4/29, I wrote perl script that could help perform the slide shortening of contigs
  • Here, I test the script/method of slide shortening to increase the contig length after recruitment
  • SAG sequence: ID9 because showed the lower contig length but showed relatively good contig length
    • Sequences found in: </media/disk-1/samchiang/Sub_tests_folder_imp/2_Extending_Mouse_Bac_SAG_contigs/BLAT_Meta2SAG_040311/Step4_pooled_together/SAG_A_ID9>
    • Test found in: </media/disk-2/cjwei/slide-shortening_Assembly_Test>
    • Used various sliding cut lengths:
      • Test 1: >1000bp contigs cut; step size = 200bp
      • Test 2: >2000bp contigs cut; step size = 100bp (same as "Whole Genome Amplification and Denovo Assembly of Single Bacterial Cells")
      • Test 3: >1000bp contigs cut; step size = 100bp
      • Test 4: >100bp contigs cut; step size = 10bp (most highly cut option)
  • Also tried out the sliding cut on best assembly (see if can improve the best recruited assembly results) = ID3
    • Tested >2000bp contigs cut; step size = 100bp (was shown with ID9 that this produced highest N50 and highest sum)

Procedure

  • Use Sliding Cut for combined SAG and recruited metagenome contigs:
perl contig_sliding_cut.pl SAG_A_ID9_and_recruted_Meta_all >SAG_A_ID9_and_recruted_Meta_all_slide_cut
  • Run Velveth
    • k-mer 27, -short
/media/disk-2/cjwei/Velvet/velveth SAG_A_ID9_and_recruted_Meta_all_slide_cut_Assembly/ 27 -fasta -short SAG_A_ID9_and_recruted_Meta_all_slide_cut
  • Run Velvetg
    • fixed setting as cov_cutoff 2, min-contig lgth 100
/media/disk-2/cjwei/Velvet/velvetg SAG_A_ID9_and_recruted_Meta_all_slide_cut_Assembly/ -cov_cutoff 2 -min_contig_lgth 100

Results

ID9 (problem assembly)

n       n:100   n:N50  min    median mean   N50    max    sum
7871    7871    737    100    196    465    1000   20981  3662057 contigs_1000_200.fa
7819     7819    710    100    197    473    1057   21081  3703868 contigs_2000_100.fa
7861    7861    726    100    196    469    1039   21081  3689257 contigs_1000_100.fa
7869    7869    718    100    196    468    1041   21091  3690448 contigs_100_10.fa
14828   14828   1140   100    170    403    902    38317  5988190 ../SAG_A_ID9_and_recruted_Meta_all
  • Based on results, notice that >2000bp contigs cut; step size = 100bp had highest N50 and sum (with relatively the same number of contigs formed -> slightly fewer may be because less short contigs?)

ID3 (best assembly)

n       n:100   n:N50  min    median mean   N50    max    sum
1818    1818    123    100    276    1270   4712   54000  2309329 contigs.fa
6232    6232    471    100    276    2816   10646  54047  17.55e6 ../SAG_A_ID3_and_recruted_Meta_all