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关于open search的几个问题 #68

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daimantianxingguangzhishi opened this issue Oct 26, 2023 · 0 comments
Open

关于open search的几个问题 #68

daimantianxingguangzhishi opened this issue Oct 26, 2023 · 0 comments

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@daimantianxingguangzhishi
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daimantianxingguangzhishi commented Oct 26, 2023

1.我们发现即使在勾选open search后,是否添加某些不常见的Variable Modification依然能够显著影响ID rate,理论上这种情况是否很少见?
如:对我们的古生物样品来说,勾选open search后,Variable Modification不添加Oxidation[P]的ID rate仅有0.3%,而添加Oxidation[P]后达到9.3%(Oxidation[P]修饰在Display All中选择)。
搜库前Open Search和Variable Modification的设置如下图所示:
test
搜库后结果对比如下图所示(上为不添加Oxidation[P],下为添加Oxidation[P]):
test
test
test
test

2.在Identification中将FDR≤1%设为Spectra水平开始搜库,搜库完成后的Summary会自动写作“FDR Less than 1% at Peptides Level”而非Spectra Level。
搜库前的设置如下图所示,设置为Spectra Level:
test
搜库完成后的Summary如下图所示,自动变为Peptide Level:
test
经我们检查认为,搜库结果确实遵循Spectra Level,那么Summary的显示是否有误?
确认过程如下:我们将FDR≤1%设为Spectra水平开始搜库,在pFind.spectra文件中含有每个PSM的q value,比较发现同一sequence相同modification的几个PSM其q value不相同,因此认为这里的q value确实是spectra层面的而非peptide层面的。在我们的数据中,从pFind.spectra中筛选q value为0.01以下的PSM,发现target有2827个,decoy有28个,pFind_Filtered.spectra报道的就是这2827个PSM,其中应该含有约28个假阳性,也就是说PSM层面的FDR为1%,搜库结果与设置是一致的。

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