Hosuk:LabNotes/2014-2-9: Difference between revisions
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====Procedure==== | ====Procedure==== | ||
*#Aligned BW images of all steps at each positions, and shift and crop the original images | |||
*#Maximum projection image with all images of 8 steps | *#Maximum projection image with all images of 8 steps | ||
*#Run PISA to make BW images | *#Run PISA to make BW images | ||
Line 80: | Line 81: | ||
====Result==== | ====Result==== | ||
=====Counts===== | =====Rolony Counts===== | ||
*'''After masking and running PISA, before decoding''' | |||
*Counts per each step were too big, I think the counts include many false positive signals. | |||
*However decoding step might filter out those false positive. | |||
[[File:RolonyCountTable.png|250px]] | |||
[[File:RolonyCount_Bar.png|600px]] | |||
=====Target Counts===== | |||
{| {{table}} border = 1 | {| {{table}} border = 1 | ||
| align="center" style="background:#f0f0f0;"|'''Target''' | | align="center" style="background:#f0f0f0;"|'''Target''' | ||
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*[[Media:DecodedEachTargetEachBinaryVariationEachPos_2014-02-09.zip|'''Dilated Images of each target, each binary variation''' ]] | *[[Media:DecodedEachTargetEachBinaryVariationEachPos_2014-02-09.zip|'''Dilated Images of each target, each binary variation''' ]] | ||
*[[Media:DecodingInfo_4.xlsx|'''Analysis Excel file''']] | *[[Media:DecodingInfo_4.xlsx|'''Analysis Excel file''']] | ||
*[[Media:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MaskedMIPofSteps.zip|'''Overlay images''']] : Red- | *[[Media:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MaskedMIPofSteps.zip|'''Overlay images''']] : (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps)) | ||
**Each target includes its 4 binary variations. | **Each target includes its 4 binary variations. | ||
*Position 1 (Red- | |||
*Position 1 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps)) | |||
**Each target includes its 4 binary variations. | **Each target includes its 4 binary variations. | ||
[[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos1.png|700px]] | [[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos1.png|700px]] | ||
*Position 2 (Red- | *Position 2 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps)) | ||
**Each target includes its 4 binary variations. | **Each target includes its 4 binary variations. | ||
[[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos2.png|700px]] | [[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos2.png|700px]] | ||
*Position 3 (Red- | *Position 3 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps)) | ||
**Each target includes its 4 binary variations. | **Each target includes its 4 binary variations. | ||
[[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos3.png|700px]] | [[File:Composite_Red-L858RMU_Green-L858RWT_Blue-T790MMU_Cyan-T790MWT_Gray-MSKD-DCDATA_Pos3.png|700px]] |
Latest revision as of 19:21, 14 February 2014
NCIH1975 : Mutation Detection : 2nd Try, Data Analysis[edit]
- Data from 2/3
- Took pictures at 3 Positions on the sample in MatTek dish
Procedure[edit]
- Aligned BW images of all steps at each positions, and shift and crop the original images
- Maximum projection image with all images of 8 steps
- Run PISA to make BW images
- Use the BW image as mask image file and do masking an each image (=image file .* mask image)
- Run PISA with masked image, and obtain a number of rolonies, centroid coordinate information of each rolony
- Decode the rolony to extract decoding information along steps
Major Parameters[edit]
Decode Binary table of each target[edit]
EGFR_T790M_WT | EGFR_T790M_MU | EGFR_L858R_WT | EGFR_L858R_MU | |
Step1 (1FAM) | 0 | 1 | 0 | 1 |
Step2 (1Cy3) | 1 | 0 | 0 | 0 |
Step3 (2FAM) | 0 | 0 | 1 | 0 |
Step4 (2Cy3) | 0 | 0 | 0 | 1 |
Step5 (3FAM) | 1 | 0 | 1 | 0 |
Step6 (3Cy3) | 0 | 1 | 0 | 0 |
Step7 (4FAM) | 1 | 0 | 0 | 1 |
Step8 (4Cy3) | 0 | 1 | 1 | 0 |
Decode Binary String of each target[edit]
- EGFR_T790M_WT = '01010010'
- EGFR_T790M_MU = '10100001'
- EGFR_L858R_WT = '10010100'
- EGFR_L858R_MU = '01001001'
Allowing 1 bit Error cases of Decode Binary String[edit]
- EGFR_T790M_WT
- Normal : '01010010'
- Error 1 : '01010000'
- Error 2 : '01000010'
- Error 3 : '00010010'
- EGFR_T790M_MU
- Normal : '10100001'
- Error 1 : '10100000'
- Error 2 : '10000001'
- Error 3 : '00100001'
- EGFR_L858R_WT
- Normal : '10010100'
- Error 1 : '10010000'
- Error 2 : '10000100'
- Error 3 : '00010100'
- EGFR_L858R_MU
- Normal : '01001001'
- Error 1 : '01001000'
- Error 2 : '01000001'
- Error 3 : '00001001'
Result[edit]
Rolony Counts[edit]
- After masking and running PISA, before decoding
- Counts per each step were too big, I think the counts include many false positive signals.
- However decoding step might filter out those false positive.
File:RolonyCountTable.png File:RolonyCount Bar.png
Target Counts[edit]
Target | Pos 1 | Pos 2 | Pos 3 | Avg | Std |
EGFR_T790M_WT | |||||
01010010 | 4 | 2 | 6 | 4 | 2 |
01010000 | 5 | 14 | 16 | 11.67 | 5.86 |
01000010 | 3 | 2 | 1 | 2 | 1 |
00010010 | 4 | 12 | 10 | 8.67 | 4.16 |
EGFR_T790M_MU | |||||
10100001 | 0 | 0 | 2 | 0.67 | 1.15 |
10100000 | 1 | 1 | 2 | 1.33 | 0.58 |
10000001 | 1 | 1 | 0 | 0.67 | 0.58 |
00100001 | 0 | 0 | 0 | 0 | 0 |
EGFR_L858R_WT | |||||
10010100 | 3 | 6 | 8 | 5.67 | 2.52 |
10010000 | 5 | 11 | 18 | 11.33 | 6.51 |
10000100 | 4 | 7 | 5 | 5.33 | 1.53 |
00010100 | 2 | 28 | 22 | 17.33 | 13.61 |
EGFR_L858R_MU | |||||
01001001 | 0 | 0 | 0 | 0 | 0 |
01001000 | 0 | 0 | 1 | 0.33 | 0.58 |
01000001 | 17 | 12 | 6 | 11.67 | 5.51 |
00001001 | 1 | 0 | 1 | 0.67 | 0.58 |
Data Files[edit]
- Data Files after Masking
- Dilated Images of each target, each binary variation
- Analysis Excel file
- Overlay images : (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps))
- Each target includes its 4 binary variations.
- Position 1 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps))
- Each target includes its 4 binary variations.
File:Composite Red-L858RMU Green-L858RWT Blue-T790MMU Cyan-T790MWT Gray-MSKD-DCDATA Pos1.png
- Position 2 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps))
- Each target includes its 4 binary variations.
File:Composite Red-L858RMU Green-L858RWT Blue-T790MMU Cyan-T790MWT Gray-MSKD-DCDATA Pos2.png
- Position 3 (Red : L858R-MU, Green : L858R-WT, Blue : T790M-MU, Cyan : T790M-WT, Gray : All Targets (Masked MIP images of all Steps))
- Each target includes its 4 binary variations.
File:Composite Red-L858RMU Green-L858RWT Blue-T790MMU Cyan-T790MWT Gray-MSKD-DCDATA Pos3.png
- Matlab code : Rolony_Decoding_v8.m