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Input data from Affymetrix arrays #10

Closed sdwien closed 4 years ago

sdwien commented 4 years ago

I tried to use the ABIS tool via the web, but got an error: "An error has occurred. Check your logs or contact the app author for clarification." It works fine for the TPMPBMC.txt file, so it is not an issue of my browser, I assume. My file contains RMA-normalized counts from human Affymetrix PrimeView arrays, has 19464 rows and 4 columns (gene symbols and 3 samples). It looks like this:

head test123.txt
"SCH1"  "SCH2"  "SCH3"
"1060P11.3" 3.5967598624897 3.93685508289532    3.87821946945781
"A1BG"  6.45565863125128    5.82575471633686    5.79043259640592
"A1CF"  5.34300130822687    5.68253584972171    5.30336643058087
"A2M"   4.15193049248871    4.3784780972333 3.94194736201775
"A2ML1" 3.49557726410993    3.6238503706137 3.51239887716308
"A3GALT2"   3.4001284612413 3.53329672413759    3.64071911619115
"A4GALT"    4.65199708894221    5.19605851776019    4.9648856544884
"A4GNT" 4.41943768837953    4.39662210867146    4.12529843617137
"AAAS"  5.73657599875531    5.8650512477877 5.86236578077812

I would be grateful for suggestions on what may be wrong here. Thanks and best regards. PS.: Are there some genes that "have to" be there? PPS.: I just noticed that when selecting "RNA-Seq" as input type, it does run and output a table, but not when selecting "Microarray". Your example file works with both settings, though, and outputs different numbers of rows with each.

giannimonaco commented 4 years ago

Hi,

I am not entirely sure why you are getting the error message only when you use microarray. The data should be on linear scale. I think that RMA-normalized counts are log2 transformed. Can you convert them back to linear scale and try again. To convert them just apply this formula to all values: 2^value

Try this first and let me know if it works.

Gianni

On Mon, 9 Mar 2020 at 12:57, sdwien notifications@github.com wrote:

I tried to use the ABIS tool via the web, but got an error: "An error has occurred. Check your logs or contact the app author for clarification." It works fine for the TPMPBMC.txt file, so it is not an issue of my browser, I assume. My file contains RMA-normalized counts from human Affymetrix PrimeView arrays, has 19464 rows and 4 columns (gene symbols and 3 samples). It looks like this:

head test123.txt "SCH1" "SCH2" "SCH3" "1060P11.3" 3.5967598624897 3.93685508289532 3.87821946945781 "A1BG" 6.45565863125128 5.82575471633686 5.79043259640592 "A1CF" 5.34300130822687 5.68253584972171 5.30336643058087 "A2M" 4.15193049248871 4.3784780972333 3.94194736201775 "A2ML1" 3.49557726410993 3.6238503706137 3.51239887716308 "A3GALT2" 3.4001284612413 3.53329672413759 3.64071911619115 "A4GALT" 4.65199708894221 5.19605851776019 4.9648856544884 "A4GNT" 4.41943768837953 4.39662210867146 4.12529843617137 "AAAS" 5.73657599875531 5.8650512477877 5.86236578077812

I would be grateful for suggestions on what may be wrong here. Thanks and best regards.

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sdwien commented 4 years ago

Hi Gianni,

thanks, I tried that. My input now looks like that:

"SCH1"        "SCH2"        "SCH3"
"1060P11.3"     12.0985300026222        15.314804898821 14.7048429344018
"A1BG"  87.7701599438797        56.7187847325014        55.346976271497
"A1CF"  40.5885615838657        51.3586668803532        39.4886578972973
"A2M"   17.7768831665421        20.7995166455123        15.3689570632719
"A2ML1" 11.279078224529 12.3278590812149        11.4113603223342
"A3GALT2"       10.5570032667953        11.5778601011023        12.4728488587772
"A4GALT"        25.1414698030673        36.6580596081672        31.2305409681272
"A4GNT" 21.3984988711742        21.0627529726873        17.4517334181841
"AAAS"  53.3189327732681        58.2849397258712        58.1765476599487

It still works when I set the input to RNA-Seq (results file attached, looks weird with numbers over 100 in some rows), but not with Microarray set where I am getting the same error as before. test123lin.txt_deconvolution.txt I can get the raw intensities for these arrays. How did you do it with the Illumina arrays you used? Thanks for your help. I would really like to implement this tool for our facility. Best, Sophia

giannimonaco commented 4 years ago

Hi Sophia,

yes, the results look weird because your data is microarray and it is not compatible with the RNA-Seq normalisation. The microarray data need to be in the linear scale and then ABIS will perform a quintile normalisation. Then you should obtain good results, although consider that you could still have some technical and biological variability.

Best, Gianni

On Tue, 10 Mar 2020 at 07:32, sdwien notifications@github.com wrote:

Hi Gianni,

thanks, I tried that. My input now looks like that:

"SCH1" "SCH2" "SCH3" "1060P11.3" 12.0985300026222 15.314804898821 14.7048429344018 "A1BG" 87.7701599438797 56.7187847325014 55.346976271497 "A1CF" 40.5885615838657 51.3586668803532 39.4886578972973 "A2M" 17.7768831665421 20.7995166455123 15.3689570632719 "A2ML1" 11.279078224529 12.3278590812149 11.4113603223342 "A3GALT2" 10.5570032667953 11.5778601011023 12.4728488587772 "A4GALT" 25.1414698030673 36.6580596081672 31.2305409681272 "A4GNT" 21.3984988711742 21.0627529726873 17.4517334181841 "AAAS" 53.3189327732681 58.2849397258712 58.1765476599487

It still works when I set the input to RNA-Seq (results file attached, looks weird with numbers over 100 in some rows), but not with Microarray set where I am getting the same error as before. test123lin.txt_deconvolution.txt https://github.com/giannimonaco/ABIS/files/4310610/test123lin.txt_deconvolution.txt I can get the raw intensities for these arrays. How did you do it with the Illumina arrays you used? Thanks for your help. I would really like to implement this tool for our facility. Best, Sophia

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