Pseudodomonas syringae

Pseudomonas syringae pv. tomato str. DC3000

Originally sequenced and finished at TIGR: published Sept 2003

Data

NCBI

 AA: no assembly
 TA 80,959 reads 
 Genome Project
 Taxonomy TaxId=223283

Chromosome + 2 plasmids:

 Name           Length    %GC
 NC_004578.1    6,397,126 58.40
 NC_004633.1    73,661    55.15
 NC_004632.1    67,473    56.17

UNC: Jeff Dangl

New sequence:

 * Solexa 3 lanes; 
 * 454 shotgun 1/4 Plate (250bp read); 
 * 454 paired ends 1/4 Plate : 
     * contain a 44 bp linker in the middle
     * the linker sequence is: GTTGGAACCGAAAGGGTTTGAATTCAAACCCTTTCGGTTCCAAC
     * there are some (not many) 454 paired end sequences that contain multiple instances of the linker (tandem): Example EUEIEUN01ANUGL_length=128_xy=0154_1891 

UNC sequence data: (not avail any more?)

 http://biology622.dhcp.unc.edu/~labweb/DCData/

UNC (e-mail):

 * Theoretical minimum number of contigs we can obtain is 268 (our reads fail to cover 269 nucleotides). 
 * Our de novo assembly spans the genome in 853 contigs totaling 6,313,026 bp. 
 * 98.7% of the genome is covered by a contig; 
 * 84% of the genome is covered by contigs 10,000 bp or greater. 
 * The average gap size between contigs is 98 bp; 
 * average contig size 7401 bp. 
 * The N50 = 37,444 bp. 
 * Our largest BAMBUS "scaffold" is 2,565,761 bp,

Data stats

 .                               #elem             min     median  max     sum             mean    stdev   n50
 DC3000.format.454Reads.fna      123,992           38      86      329     15623908        126.01  58.89   142     DC3000 Paired End Reads (forward+linkerr+reverse)
 DC3000.TCA.454reads.format.fna  77,466            35      244     371     18627363        240.46  26.85   245     DC3000 454 Reads
 DC3000.reads.filtered.fasta     6,340,136         32      32      32      202884352       32      0       32      DC3000 Solexa Reads
 DC3000Plasmids.fa               2                 67473   73661   73661   141134          70567   3094    73661   Pseudomonas syringae pv. tomato DC3000 Plasmids
 Psudomonas_syringae.fa          1                 6397126 6397126 6397126 6397126         6397126 0       6397126 Pseudomonas syringae pv. tomato DC3000 reference
 
 
 Quality values are missing for all data sets!!!
 I assigned default qual=3 to all the base (.frg & .afg files)  

Files location:

 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Data
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly

Assemblies

CBCB

1. AMOSCmp

 454 single reads + Solexa reads 
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Solexa-454/2007_1009_AMOSCmp-relaxed
 142 contigs (37 negative gaps, 89 positive gaps)
 No read trimming was done. 
 AMOScmp used the following parameters:
   nucmer -c  20
   casm-layout -t 20 -o 5
 "-t 20" allows for 20 bp long  dirty sequence ends which seem to solve the "low quality" problem.
 => 22 large contigs
 
 454 single reads + 30 bp Solexa reads  => 167 contigs , 49 negative gaps, 100 positive gaps 
 454 single reads + 25 bp Solexa reads  => 293 contigs,  144 negative gaps, 131 positive gaps 

2. AMOSCmp

 454 single reads + Solexa reads + 454 paired ends
 Only the 454 paired ends that contain 1 single complete adaptor sequence were used (allmost all)
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Solexa-454-454p/2007_1011_AMOSCmp-relaxed-filtered
 149 contigs; very similar to the prev ome

3. AMOSCmp (MAJORITY=50)

 454 single reads + Solexa reads 
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Solexa-454/2007_1015_AMOSCmp-relaxed-MAJORITY50 
 131 contigs  (18 negative gaps)
 No read trimming was done. 
 AMOScmp used the following parameters:
   nucmer -c  20
   casm-layout -t 20 -o 5 -m 50
 No read trimming was done. 
 "-t 20" allows for 20 bp long  dirty sequence ends which seem to solve the "low quality" problem.
 "-m 20" merges some contigs together
 => 10 large contigs

 contig#        len     gc%
 4              2290968 59.00
 7              1817904 58.18
 3              1405326 58.08
 5              648413  58.48
 2              192413  57.86
 6              87152   58.02
 131            71251   56.47
 1              32939   54.86
 130            29120   59.36
 9              20309   53.56
 95             3589    59.46

4. AMOSCmp

 Sanger reads
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Sanger/2007_1011_AMOSCmp-relaxed
 Many miss-oriented mates in the 4.8M-5M region of the chromosome
 22 contigs
 Chromosome
 Chromosome problem

5. Celera 3.11

 Sanger reads
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Sanger/2007_1011_WGA
 22 scaff, 46 contigs, 181 degens
 Scaffold 7180000001443 looks circular: possible 163,074 bp plasmid
 aligns to 4.8M-5M "problem" region in the chromosome
 7180000001443.png

     [S1]     [E1]  |     [S2]     [E2]  |  [LEN 1]  [LEN 2]  |  [% IDY]  |  [LEN R]  [LEN Q]  |  [COV R]  [COV Q]  | [TAGS]
 ===============================================================================================================================
        1   175592  |        1   175592  |   175592   175592  |   100.00  |   175592   175592  |   100.00   100.00  | 7180000001443   7180000001443   [IDENTITY]
        1    12519  |   163075   175592  |    12519    12518  |    99.98  |   175592   175592  |     7.13     7.13  | 7180000001443   7180000001443   [BEGIN]
   163075   175592  |        1    12519  |    12518    12519  |    99.98  |   175592   175592  |     7.13     7.13  | 7180000001443   7180000001443   [END]

     [S1]     [E1]  |     [S2]     [E2]  |  [LEN 1]  [LEN 2]  |  [% IDY]  |  [LEN R]  [LEN Q]  |  [COV R]  [COV Q] | [TAGS]
 ===============================================================================================================================
  4790727  4911492  |   120764        1  |   120766   120764  |    99.98  |  6397126   175592  |     1.89    68.78  | gi|28867243|ref|NC_004578.1|    7180000001443
  4898971  4955870  |   175592   118697  |    56900    56896  |    99.98  |  6397126   175592  |     0.89    32.40  | gi|28867243|ref|NC_004578.1|    7180000001443

6. AMOSCmp (Chromosome+3 plasmids ref)

 Sanger reads
 Reference=complete genome(chromosome+3 plasmids) use "circular contig" in Celera 3.11 assembly
 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Sanger/2007_1012_AMOSCmp-relaxed-3plasmids
 38 contigs: 15 for main chromosome, 1 for longer plasmid, 21 for shorter plasmid, 1 for "circular contig"
 The missoriented read pile corresponding to the chromosome (4. AMOSCmp of Sanger reads) has dissapeared
 AA ready for submission: /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Sanger/2007_1012_AMOSCmp-relaxed-3plasmids/AA/umd-20071030-141700.tar.gz

Solexa assemblied for different read coverages

Location

 /fs/szasmg2/Bacteria/Pseudodomonas_syringae/Assembly/Solexa/sample/
 
 Several AMOScmp assemblies, using 100%, 90% ... 10% of the P. syringae Solexa reads. These would correspond to 30X, 27X, 24X .. 3X coverage 
 The read sampling was done randomly. One sample set for each coverage.

Chromosome + 2 plasmids

qc stats for Solexa assemblies done at different coverage levels

 cvg: 30,27,24...3
   
 $ more contig.chromo.summary positiveGaps.chromo.summary
 ::::::::::::::
 contig.summary
 ::::::::::::::
 %reads  #elem   #elem0  #elem<0 min     median  max     sum     mean    stdev   n50
 100     5502    0       0       32      338     32148   7296600 1326.17 2157.6  3714
 90      6463    0       0       32      330     25252   7252304 1122.13 1799.43 3009
 80      7570    0       0       32      303     20690   7209479 952.38  1487.03 2573
 70      9030    0       0       32      309     26306   7170384 794.06  1219.53 1986
 60      10571   0       0       32      295     22249   7124996 674.01  961.22  1608
 50      12598   0       0       32      274     22204   7075934 561.67  767.55  1266
 40      15343   0       0       32      252     9176    7011485 456.98  575.64  934
 30      21248   0       0       32      202     7751    6931907 326.24  376.06  597
 20      38702   0       0       32      117     3276    6807914 175.91  178.92  278
 10      84545   0       0       32      56      2652    6267925 74.14   57.62   90
 ::::::::::::::
 positiveGaps.summary
 ::::::::::::::
 %reads  #elem   #elem0  #elem<0 min     median  max     sum     mean    stdev   n50
 100     117     10      0       0       22      3065    19625   167.74  418.75  1308
 90      130     16      0       0       19      2100    19725   151.73  369.07  1211
 80      142     18      0       0       15      2174    20034   141.08  361.86  1209
 70      178     15      0       0       9       3417    20443   114.85  395.13  1823
 60      263     35      0       0       6       3875    21161   80.46   345.97  1457
 50      450     64      0       0       4       3398    22305   49.57   278.39  1823
 40      1047    156     0       0       4       3398    26488   25.3    173.77  929
 30      2915    446     0       0       4       3426    39094   13.41   115.74  104
 20      11154   1324    0       0       5       3420    110485  9.91    57.22   19
 10      44751   3321    0       0       9       3875    631930  14.12   35.45   25

Chromosome (only)

 $ more contig.chromo.summary positiveGaps.chromo.summary
 ::::::::::::::
 contig.chromo.summary
 ::::::::::::::
 %reads  #elem   #elem0  #elem<0 min     median  max     sum     mean    stdev   n50
 100     5352    0       0       32      387     18942   7152892 1336.49 2069.25 3674
 90      6313    0       0       32      362     16470   7110882 1126.39 1721.34 2969
 80      7411    0       0       32      322     15227   7069778 953.96  1436.49 2521
 70      8865    0       0       32      324     14901   7032202 793.25  1154.9  1968
 60      10406   0       0       32      304     10231   6988498 671.58  919.5   1586
 50      12389   0       0       32      279     7246    6941706 560.31  733.75  1247
 40      15131   0       0       32      256     5409    6879810 454.68  554.17  920
 30      20998   0       0       32      204     4102    6801160 323.9   358.93  588
 20      38368   0       0       32      117     2220    6680303 174.11  170.14  274
 10      83839   0       0       32      56      762     6144687 73.29   51.16   89
 ::::::::::::::
 positiveGaps.chromo.summary
 ::::::::::::::
 .       #elem   #elem0  #elem<0 min     median  max     sum     mean    stdev   n50
 100     15      5       0       0       1       33      107     7.13    10.84   33
 90      24      7       0       0       2       42      146     6.08    10.38   42
 80      38      11      0       0       2       36      212     5.58    8.84    26
 70      76      11      0       0       3       33      413     5.43    6.66    11
 60      163     29      0       0       4       33      1016    6.23    7.04    13
 50      347     60      0       0       3       49      1843    5.31    6.45    11
 40      947     151     0       0       4       53      5709    6.03    7.18    12
 30      2819    442     0       0       4       63      17882   6.34    7.34    12
 20      11029   1320    0       0       5       610     88516   8.03    10.84   15
 10      44485   3313    0       0       9       197     606841  13.64   15.05   24