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wososa / PSI-Sigma

PSI-Sigma
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interpretting output results #45

Closed parsboy66 closed 1 year ago

parsboy66 commented 1 year ago

Hi there, nice job! well done.

actually, I ran the pipeline, seems works well, so I expect the output according to --fmod 0 , to have delta-PSI > 10% and p-value < 0.01.

but at the output for the whole results, T-test Pvalue and FDR are 1.0000. ex: chr10:90894892-90895879 RPP30 chr10:90895454-90895483 SES 1 0 - TCONS.00002309 -94.00 1.00000e+00 1.00000e+00 94.2857142857143 na chr10_90894892_90895879_W_TCONS.00002309_1 chr1:156592671-156593463 NAXE chr1:156593408-156593463 A3SS 1 0 - TCONS.00000182 -95.00 1.00000e+00 1.00000e+00 95 na chr1_156592671_156593463_S_TCONS.00000182_1 chr19:35745701-35745872 PSENEN chr19:35745835-35745872 TSS|A3SS 1 0 - TSS.TCONS.00010534 -95.00 1.00000e+00 1.00000e+00 95 na chr19_35745701_35745872_S_TSS.TCONS.00010534_1 chr19:10679928-10681018 ILF3 chr19:10680346-10680443 SES 1 0 - TCONS.00010462 -95.00 1.00000e+00 1.00000e+00 95 na chr19_10679928_10681018_W_TCONS.00010462_1 chr1:44724393-44725135 ARMH1 chr1:44724762-44724839 MES 1 0 - TCONS.00000089 -100.00 1.00000e+00 1.00000e+00 100 na chr1_44724393_44725135_W_TCONS.00000089_2 chr1:44724393-44725135 ARMH1 chr1:44724539-44724668 MES 1 0 - TCONS.00000089 -100.00 1.00000e+00 1.00000e+00 100 na chr1_44724393_44725135_W_TCONS.00000089_1 chr1:229271371-229295847 RAB4A chr1:229286486-229286566 MES 1 0 - TCONS.00000272 -100.00 1.00000e+00 1.00000e+00 100 na chr1_229271371_229295847_W_TCONS.00000272_1

do you have any suggestions to interpret it?

also, another question is, which configuration is the best for low-depth (coverage) samples?

Thanks

wososa commented 1 year ago

Hi @parsboy66 ,

Sorry for the late reply. When p-value equals 1, it means that the number of "valid" replicates in groupa.txt and groupb.txt is too low to get a p-value. For example, in your case, the number of control samples equal 1, it is below the minimal number of p-value calculation. To get a p-value, you will need to put more samples in groupa.txt and groupb.txt files.

It is always challenging to analyze low-depth samples. The delta-PSI > 10% and p-value < 0.01 criteria can be too very stringent for low-depth samples because a splicing event can have no reads in most of the samples in groupa.txt or groupb.txt. If you can't increase the number of replicates, you will need to make sure that the splicing events of your interest are reproducible in other datasets. If an exon-skipping event is reproducible in a second dataset, the splicing event is likely to be real. The best would be doing wet-lab experiments (e.g., RT-PCR) to validate it.

Let me know if you need further explanations.

Best, Woody

parsboy66 commented 1 year ago

Dear Woody

yeap you are right, the number of samples was the problem which is solved. just another naive question about the outout interpretation: N and T indicate the number of samples supporting the event from groupa.txt(N) and groupb.txt (T). dPSI also is the PSI(N) - PSI(T) ? and if so, positive dPSI shows the event in groupa(N) while the negative dPSI shows the active events in groupb(T). is it correct the correct way to interpret the results? how about N and T values?

Thanks in advance

wososa commented 1 year ago

Hi @parsboy66 ,

N, T, N value, and T value are explained in Readme. dPSI means the difference of ‘PSI’ values (groupb average - groupa average). For example, positive dPSI means exon-inclusion in samples of groupb.txt. Negative dPSI means exon-skipping in samples of groupb.txt.

Woody

parsboy66 commented 1 year ago

Thanks woody, clear.

On Wed, 10 May 2023, 13:44 Woody Lin, @.***> wrote:

Hi @parsboy66 https://github.com/parsboy66 ,

N, T, N value, and T value are explained in Readme. dPSI means the difference of ‘PSI’ values (groupb average - groupa average). For example, positive dPSI means exon-inclusion in samples of groupb.txt. Negative dPSI means exon-skipping in samples of groupb.txt.

Woody

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