Sam:LabNotes/Microbiome-new/2011-2-15: Difference between revisions
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>Sam Chiang |
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==Discussions== |
Revision as of 02:37, 28 February 2011
Bowtie mapping(whole reads) and contamination analysis(failed to aligned reads)- HC-MDA-Ecoli-SAGs(NX1-NX8)-Jan09
Background
- Ecoli SAGs were prepared from flow-sorted single E.coli cells.
- Eight E.coli SAGs samples were made to Nextera-libraries.
- These libraries were equally pulled (20ng per library) and sequenced in pair-end(PE) mode.
- The sequenced data processed by Dr. Zhang for base-calling and saved into FASTQ files.
Wiki(12-29-'10) Sample no. Sample name Total DNA(ng) ----------------------------------------------- NX01 WGA102710-(1-4)- 3hr 288.50ng NX02 WGA102710-(1-4)-10hr 500.00ng NX03 WGA110310-(5)- 3hr 114.50ng NX04 WGA110310-(5)- 10hr 500.00ng
Wiki(01-06-'11) Sample no. Sample name Total DNA(ng) ----------------------------------------------- NX05 WGA122010-(1-2)-3hr 181.50ng NX06 WGA122010-(1-2)-10hr 200.00ng NX07 WGA122110-(2-3)-3hr 50.00ng NX08 WGA122110-(2-3)-10hr 200.00ng
Analysis approaches
Data pooling
- Using "gunzip" command to unzip SE files.
- Combine two SE data(read1 + read2) into one PE data
e.g. samchiang@genemapster:~/Raw-Read-backup/110120_HL087$ cat s_4_1_ID8.txt s_4_2_ID8.txt > s_4_pe_ID8.txt &
Genome coverage analysis (whole reads and 1000000 down-sampled reads
- Script: Read2coverage-kz-3
- Data resource: pooled pe reads data from HC-MDA-Ecoli-SAGs(NX1-NX8)-Jan09
- Command
perl Read2coverage-kz-3 /media/disk-2/samchiang/Raw-Read-backup/110120_HL087/pooled_pe/s_4_pe_ID8_3074085.txt /media/disk-2/samchiang/Tools/bowtie-0.12.5/indexes/e_coli_k12_mg1655/e_coli_k12_mg1655 &
- Bowtie setting
samchiang@genemapster:~/Scripts$ /media/disk-2/samchiang/Tools/bowtie-0.12.5/bowtie -k 1 -l 28 -n 2 -m 1 --best --best --phred64-quals
Results
- Original excel table Media:021511_whole-read-bowtie-NX1-8.xls
Sequence Read Name | s_4_pe_ID1_NX | s_4_pe_ID2_NX | s_4_pe_ID3_NX | s_4_pe_ID4_NX | s_4_pe_ID5_NX | s_4_pe_ID6_NX | s_4_pe_ID7_NX | s_4_pe_ID8_NX |
Library Nick Name | NX01 | NX02 | NX03 | NX04 | NX05 | NX06 | NX07 | NX08 |
Library Sample Name | WGA102710_(1_4)_3hr | WGA102710_(1_4)_10hr | WGA102710_(5)_3hr | WGA102710_(5)_10hr | WGA122010_(1-2)_3hr | WGA122010_(1-2)_10hr | WGA122010_(2-3)_3hr | WGA122010_(2-3)_10hr |
DNA starting amount (ng) | 288.5 | 500.0 | 114.5 | 500.0 | 181.5 | 200.0 | 50.0 | 200.0 |
Proccessed reads | 13395058 | 8843072 | 6451756 | 15366518 | 5071132 | 7353034 | 6647566 | 4888674 |
Mapped read, no. (unique) | 2707193 (20.21%) | 5734751 (64.85%) | 1988199 (30.82%) | 8571871 (55.78%) | 742103 (14.63%) | 2963238 (40.30%) | 1048466 (15.77%) | 1590418 (32.53%) |
Failed to align, no. | 10515640 (78.50%) | 2512119 (28.41%) | 4285220 (66.42%) | 4736204 (30.82%) | 4311962 (85.03%) | 4338695 (59.01%) | 5549873 (83.49%) | 3176843 (64.98%) |
Mapped read, no (non-unique) | 172225 (1.29%) | 596202 (6.74%) | 178337 (2.76%) | 2058443 (13.40%) | 17067 (0.34%) | 51101 (0.69%) | 49227 (0.74%) | 121413 (2.48%) |
Total bps coverred, bp | 2146219 | 2001793 | 797791 | 835527 | 914841 | 767267 | 1282002 | 738538 |
Genome coverage | 46.26% | 43.15% | 17.20% | 18.01% | 19.72% | 16.54% | 27.63% | 15.92% |
Sequence Read Name | s_4_pe_ID1_NX | s_4_pe_ID2_NX | s_4_pe_ID3_NX | s_4_pe_ID4_NX | s_4_pe_ID5_NX | s_4_pe_ID6_NX | s_4_pe_ID7_NX | s_4_pe_ID8_NX |
1000000 uniqe reads | 4948046 | 1542020 | 3244646 | 1792757 | 6835270 | 2481390 | 6341154 | 3074085 |
700000 uniqe reads | 4784689 | 1736973 | ||||||
Proccessed reads | 4948046 | 1542020 | 3244646 | 1792757 | 4784689 | 1736973 | 6341154 | 3074085 |
Mapped read, no. (unique) | 1028262 (20.78%) | 1047304 (67.92%) | 1032729 (31.83%) | 1060752 (59.17%) | 701069 (14.65%) | 721899 (41.56%) | 1001553 (15.79%) | 1029838 (33.50%) |
Failed to align, no. | 3853931 (77.89%) | 381914 (24.77%) | 2118408 (65.29%) | 467033 (26.05%) | 4067507 (85.01%) | 1002826 (57.73%) | 5292536 (83.46%) | 1965747 (63.95%) |
Mapped read, no (non-unique) | 65853 (1.33%) | 112802 (7.32%) | 93509 (2.88%) | 264972 (14.78%) | 16113 (0.34%) | 12248 (0.71%) | 47065 (0.74%) | 78500 (2.55%) |
Total bps coverred, bp | 1890835 | 1445489 | 652120 | 310201 | 897507 | 456971 | 1267625 | 676415 |
Genome coverage | 40.76% | 31.16% | 14.06% | 6.69% | 19.35% | 9.85% | 27.32% | 14.58% |