评估测序文库复杂度 Library Complexity

  • A+
所属分类:Epigenetics

在制备测序文库时,经常会增加PCR的步骤来扩增DNA片段。如何评估PCR的效果和影响,本文主要分享ENCODE中针对ChIP-Seq和ATAC-Seq标准来说明。

ENCODE中主要通过三个参数来反应Library Complexity:PCB1PBC2和NRF。以下分别介绍各自的定义:

PCR Bottlenecking Coefficient 1 (PBC1)

  • PBC1=M1/M_DISTINCT where
    • M1: number of genomic locations where exactly one read maps uniquely
    • M_DISTINCT: number of distinct genomic locations to which some read maps uniquely

 

PCR Bottlenecking Coefficient 2 (PBC2)

  • PBC2= M1/M2 where
    • M1: number of genomic locations where only one read maps uniquely
    • M2: number of genomic locations where two reads map uniquely

 

Non-Redundant Fraction (NRF) - Number of distinct uniquely mapping reads (i.e. after removing duplicates) / Total number of reads.

 

如何计算这三个数值呢,在我们拿到比对的结果后(SAM或者BAM文件)

对于single end测序,以align.bam为例:

对于pair-end测序,以align.bam为例:

最后文件中包含7列,分别为:

1)TotalReadPairs

2)DistinctReadPairs

3)OneReadPair

4)TwoReadPairs

5)NRF=Distinct/Total

6)PBC1=OnePair/Distinct

7)PBC2=OnePair/TwoPair

 

对于ChIP-seq结果解读:

评估测序文库复杂度 Library Complexity

对于ATAC-Seq结果解读:

评估测序文库复杂度 Library Complexity

发表评论

:?: :razz: :sad: :evil: :!: :smile: :oops: :grin: :eek: :shock: :???: :cool: :lol: :mad: :twisted: :roll: :wink: :idea: :arrow: :neutral: :cry: :mrgreen:

目前评论:1   其中:访客  1   博主  0

    • 请输入您的QQ号 请输入您的QQ号 0

      求一个邀请码,我想在plob注册!!