Rui:RNAseq analysis on HL109: Difference between revisions
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less /home/ruiliu/SeqStore/111112_HL109/*_Indx96.txt > Indx96.txt | less /home/ruiliu/SeqStore/111112_HL109/*_Indx96.txt > Indx96.txt | ||
===Tophat_11.20.11=== | ===Tophat_11.20.11 Mm folder=== | ||
* Using UCSC bowtieIndex and Annotation | * Using UCSC bowtieIndex and Annotation | ||
* "tophat" batch command | * "tophat" batch command | ||
Line 27: | Line 27: | ||
tophat -p 10 --solexa1.3-quals -o Indx95 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx95.txt | tophat -p 10 --solexa1.3-quals -o Indx95 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx95.txt | ||
tophat -p 10 --solexa1.3-quals -o Indx96 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx96.txt | tophat -p 10 --solexa1.3-quals -o Indx96 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx96.txt | ||
===Tophat_11.20.11=== | |||
====Re-download UCSC_Hs package from iGenome==== | |||
wget ftp://igenome:G3nom3s4u@ftp.illumina.com/Homo_sapiens/UCSC/hg19/Homo_sapiens_UCSC_hg19.tar.gz | |||
tar -zxvf Homo_sapiens_UCSC_hg19.tar.gz | |||
BowtieIndex: ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome | |||
Annotation: ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf | |||
====Re-run tophat==== | |||
tophat -p 10 --solexa1.3-quals -o Indx88 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx88.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx89 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx89.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx90 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx90.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx91 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx91.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx92 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx92.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx93 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx93.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx94 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx94.txt | |||
tophat -p 10 --solexa1.3-quals -o Indx95 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx95.txt | |||
====Stat.==== | |||
* Mapping rate is lower than previous RNAseq library (60-70%) | |||
* Samples Indx95 and 96 has severe contamination of mouse genome | |||
{| {{table}} border=1 | |||
| align="center" style="background:#f0f0f0;"|'''Hg19''' | |||
| align="center" style="background:#f0f0f0;"|'''bowtie.log''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx88''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx89''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx90''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx91''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx92''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx93''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx94''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx95''' | |||
| align="center" style="background:#f0f0f0;"|'''Indx96''' | |||
|- | |||
| ||processed||10,823,774||12,545,719||16,917,879||16,739,939||14,439,641||12,441,496||17,775,293||15,850,762||4,850,658 | |||
|- | |||
| ||one alignment||6,030,520||7,206,915||9,344,109||9,210,627||8,199,746||7,211,466||8,315,444||6,324,495||660,512 | |||
|- | |||
| ||||55.72%||57.45%||55.23%||55.02%||56.79%||57.96%||46.78%||39.90%||13.62% | |||
|- | |||
| ||failed||4,761,793||5,293,227||7,540,575||7,511,226||6,224,057||5,203,180||9,424,809||9,498,774||4,187,420 | |||
|- | |||
| ||||43.99%||42.19%||44.57%||44.87%||43.10%||41.82%||53.02%||59.93%||86.33% | |||
|- | |||
| ||suppressed -m||31,461||45,577||33,195||18,086||15,838||26,850||35,040||27,493||2,726 | |||
|- | |||
| ||||0.29%||0.36%||0.20%||0.11%||0.11%||0.22%||0.20%||0.17%||0.06% | |||
|- | |||
| |||||||||||||||||||| | |||
|- | |||
| Mm9||processed||10,823,774||12,545,719||16,917,879||16,739,939||14,439,641||12,441,496||17,775,293||15,850,762||4,850,658 | |||
|- | |||
| ||one alignment||83,419||97,933||126,687||126,258||114,051||88,122||1,442,690||3,232,448||2,099,460 | |||
|- | |||
| ||||0.77%||0.78%||0.75%||0.75%||0.79%||0.71%||8.12%||20.39%||43.28% | |||
|- | |||
| ||failed||10,738,536||12,445,311||16,788,581||16,611,039||14,322,796||12,351,258||16,322,399||12,591,421||2,734,829 | |||
|- | |||
| ||||99.21%||99.20%||99.24%||99.23%||99.19%||99.27%||91.83%||79.44%||56.38% | |||
|- | |||
| ||suppressed -m||1,819||2,475||2,611||2,642||2,794||2,116||10,204||26,893||16,369 | |||
|- | |||
| ||||0.02%||0.02%||0.02%||0.02%||0.02%||0.02%||0.06%||0.17%||0.34% | |||
|} | |||
===Cuffdiff_11.22.11=== | |||
* comparison for Wei: [http://genome-tech.ucsd.edu/LabNotes/index.php/Rui_Liu#HL106.2FHL109] | |||
* uncertainty about the #4 and #5, b/c separate pair-wise comparisons showed different results/DE gene # from combined all three in one cuffdiff | |||
# Indx88:91 - Ctrl/0d vs Ctrl/2d | |||
# Indx89:92 - B1/0d vs B1/2d | |||
# Indx90:93 - B3/0d vs B3/2d | |||
# Indx88:89:90 - Ctrl/0d vs B1/0d vs B3/0d | |||
# Indx91:92:93 - Ctrl/2d vs B1/2d vs B3/2d | |||
* comparison for Dr. Xu | |||
# Indx94:95 - Ctrl vs RFP | |||
# Indx94:96 - Ctrl vs RFP/GFP | |||
# Indx95:96 - RFP vs GFP/RFP | |||
===Tophat_11.28.11=== | |||
* Re-run tophat against Ensemble Human dataset | |||
===Cuffdiff_11.28.11=== | |||
* Re-run cuffdiff against Ensemble Human dataset |
Latest revision as of 00:06, 1 December 2011
HL109[edit]
- Samples: assumed to be HL106 [1]
Data[edit]
- Transfer files to Indx_seq folder, save as "less" batch command, Indx92,95 and 96 were also used by Noi, repeat less
less /home/ruiliu/SeqStore/111112_HL109/*_Indx88.txt > Indx88.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx89.txt > Indx89.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx90.txt > Indx90.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx91.txt > Indx91.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx92.txt > Indx92.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx93.txt > Indx93.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx94.txt > Indx94.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx95.txt > Indx95.txt less /home/ruiliu/SeqStore/111112_HL109/*_Indx96.txt > Indx96.txt
Tophat_11.20.11 Mm folder[edit]
- Using UCSC bowtieIndex and Annotation
- "tophat" batch command
tophat -p 10 --solexa1.3-quals -o Indx88 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx88.txt tophat -p 10 --solexa1.3-quals -o Indx89 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx89.txt tophat -p 10 --solexa1.3-quals -o Indx90 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx90.txt tophat -p 10 --solexa1.3-quals -o Indx91 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx91.txt tophat -p 10 --solexa1.3-quals -o Indx92 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx92.txt tophat -p 10 --solexa1.3-quals -o Indx93 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx93.txt tophat -p 10 --solexa1.3-quals -o Indx94 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx94.txt tophat -p 10 --solexa1.3-quals -o Indx95 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx95.txt tophat -p 10 --solexa1.3-quals -o Indx96 -G ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/Mus_musculus/UCSC/mm9/Sequence/BowtieIndex/genome ../Indx_seq/Indx96.txt
Tophat_11.20.11[edit]
Re-download UCSC_Hs package from iGenome[edit]
wget ftp://igenome:G3nom3s4u@ftp.illumina.com/Homo_sapiens/UCSC/hg19/Homo_sapiens_UCSC_hg19.tar.gz tar -zxvf Homo_sapiens_UCSC_hg19.tar.gz
BowtieIndex: ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome Annotation: ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf
Re-run tophat[edit]
tophat -p 10 --solexa1.3-quals -o Indx88 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx88.txt tophat -p 10 --solexa1.3-quals -o Indx89 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx89.txt tophat -p 10 --solexa1.3-quals -o Indx90 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx90.txt tophat -p 10 --solexa1.3-quals -o Indx91 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx91.txt tophat -p 10 --solexa1.3-quals -o Indx92 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx92.txt tophat -p 10 --solexa1.3-quals -o Indx93 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx93.txt tophat -p 10 --solexa1.3-quals -o Indx94 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx94.txt tophat -p 10 --solexa1.3-quals -o Indx95 -G ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Annotation/Genes/genes.gtf ~/RNAseq/iGenome/UCSC_hg19/Homo_sapiens/UCSC/hg19/Sequence/BowtieIndex/genome ../Indx_seq/Indx95.txt
Stat.[edit]
- Mapping rate is lower than previous RNAseq library (60-70%)
- Samples Indx95 and 96 has severe contamination of mouse genome
Hg19 | bowtie.log | Indx88 | Indx89 | Indx90 | Indx91 | Indx92 | Indx93 | Indx94 | Indx95 | Indx96 |
processed | 10,823,774 | 12,545,719 | 16,917,879 | 16,739,939 | 14,439,641 | 12,441,496 | 17,775,293 | 15,850,762 | 4,850,658 | |
one alignment | 6,030,520 | 7,206,915 | 9,344,109 | 9,210,627 | 8,199,746 | 7,211,466 | 8,315,444 | 6,324,495 | 660,512 | |
55.72% | 57.45% | 55.23% | 55.02% | 56.79% | 57.96% | 46.78% | 39.90% | 13.62% | ||
failed | 4,761,793 | 5,293,227 | 7,540,575 | 7,511,226 | 6,224,057 | 5,203,180 | 9,424,809 | 9,498,774 | 4,187,420 | |
43.99% | 42.19% | 44.57% | 44.87% | 43.10% | 41.82% | 53.02% | 59.93% | 86.33% | ||
suppressed -m | 31,461 | 45,577 | 33,195 | 18,086 | 15,838 | 26,850 | 35,040 | 27,493 | 2,726 | |
0.29% | 0.36% | 0.20% | 0.11% | 0.11% | 0.22% | 0.20% | 0.17% | 0.06% | ||
Mm9 | processed | 10,823,774 | 12,545,719 | 16,917,879 | 16,739,939 | 14,439,641 | 12,441,496 | 17,775,293 | 15,850,762 | 4,850,658 |
one alignment | 83,419 | 97,933 | 126,687 | 126,258 | 114,051 | 88,122 | 1,442,690 | 3,232,448 | 2,099,460 | |
0.77% | 0.78% | 0.75% | 0.75% | 0.79% | 0.71% | 8.12% | 20.39% | 43.28% | ||
failed | 10,738,536 | 12,445,311 | 16,788,581 | 16,611,039 | 14,322,796 | 12,351,258 | 16,322,399 | 12,591,421 | 2,734,829 | |
99.21% | 99.20% | 99.24% | 99.23% | 99.19% | 99.27% | 91.83% | 79.44% | 56.38% | ||
suppressed -m | 1,819 | 2,475 | 2,611 | 2,642 | 2,794 | 2,116 | 10,204 | 26,893 | 16,369 | |
0.02% | 0.02% | 0.02% | 0.02% | 0.02% | 0.02% | 0.06% | 0.17% | 0.34% |
Cuffdiff_11.22.11[edit]
- comparison for Wei: [2]
- uncertainty about the #4 and #5, b/c separate pair-wise comparisons showed different results/DE gene # from combined all three in one cuffdiff
- Indx88:91 - Ctrl/0d vs Ctrl/2d
- Indx89:92 - B1/0d vs B1/2d
- Indx90:93 - B3/0d vs B3/2d
- Indx88:89:90 - Ctrl/0d vs B1/0d vs B3/0d
- Indx91:92:93 - Ctrl/2d vs B1/2d vs B3/2d
- comparison for Dr. Xu
- Indx94:95 - Ctrl vs RFP
- Indx94:96 - Ctrl vs RFP/GFP
- Indx95:96 - RFP vs GFP/RFP
Tophat_11.28.11[edit]
- Re-run tophat against Ensemble Human dataset
Cuffdiff_11.28.11[edit]
- Re-run cuffdiff against Ensemble Human dataset