Model-based Analysis of Genome-wide CRISPR-Cas9 Knockout (MAGeCK_NEST) is a computational tool to identify important genes from the recent genome-scale CRISPR-Cas9 knockout screens technology. Based on the previous generation, MAGeCK_VISPR, MAGeCK_NEST includes some new features, including:
* Outliers removal.
* Protein-protein interaction information integration.
* Histogram of beta scores (modified log fold changes)
* QQ-plot of p-values
* Gene set enrichment analysis (GSEA) using positive control genes as quality control
The input of the MAGeCK-NEST workflow are read count files. The formats are sgRNA, gene, readcounts Ex:
sgRNA gene sample1_readcount sample2_readcount...
gene1_gRNA_1 gene1 557 421
gene1_gRNA_2 gene1 295 128
. . .
. . .
. . .
gene2_gRNA_1 gene2 173 68
gene2_gRNA_2 gene2 85 38
. . .
. . .
. . .
mageck_nest.py nest [-h] [-n OUTPUT_PREFIX]
[--genes-varmodeling GENES_VARMODELING]
[-i INCLUDE_SAMPLES] [-b BETA_LABELS]
[--adjust-method {fdr,holm,pounds}] [-o] [-p]
[--norm-method {none,median,total,control}]
[-e NEGATIVE_CONTROL]
-k COUNT_TABLE
-d DESIGN_MATRIX
optional arguments:
-h, --help show this help message and exit
-n OUTPUT_PREFIX, --output-prefix OUTPUT_PREFIX
The prefix of the output file(s).
--genes-varmodeling GENES_VARMODELING
The number of genes for mean-variance modeling.
Default all.
-i INCLUDE_SAMPLES, --include-samples INCLUDE_SAMPLES
Specify the sample labels if the design matrix is not
given by file in the --design-matrix option. Sample
labels are separated by ",", and must match the labels
in the count table.
-b BETA_LABELS, --beta-labels BETA_LABELS
Specify the labels of the variables (i.e., beta), if
the design matrix is not given by file in the
--design-matrix option. Should be separated by ",",
and the number of labels must equal to (# columns of
design matrix), including baseline labels. Default
value: "bata_0,beta_1,beta_2,...".
--adjust-method {fdr,holm,pounds}
Method for sgrna-level p-value adjustment, including
false discovery rate (fdr), holm's method (holm), or
pounds's method (pounds).
-o, --outliers_removal
Speicify whehter you want to remove outliers and
recalculate..
-p, --PPI_prior Specify whether you want to incorporate PPI as prior
--norm-method {none,median,total,control}
Method for normalization, including "none"
(nonormalization), "median" (median normalization,
default), "total" (normalization by total read
counts), "control" (normalization by control sgRNAs
specified by the --control-sgrna option).
-e NEGATIVE_CONTROL, --negative_control NEGATIVE_CONTROL
The gene name of negative controls.
-k COUNT_TABLE, --count_table COUNT_TABLE
Provide a tab-separated count table. Each line in the
table should include sgRNA name (1st column), target
gene (2nd column) and read counts in each sample.
-d DESIGN_MATRIX, --design_matrix DESIGN_MATRIX
Provide a design matrix, either a file name or a
quoted string of the design matrix. For example,
"1,1;1,0". The row of the design matrix must match the
order of the samples in the count table (if --include-
samples is not specified), or the order of the samples
by the --include-samples option.
- gene_summary.txt
Gene: gene name
sgRNA: number of targeting sgRNA
beta_1|beta: beta score
beta_1|z: z-statistic of beta score
beta_1|wald-p-value: p-value
beta_1|wald-fdr: false discovery rate
- sgrna_summary.txt
Gene: gene name
sgRNA: sgrna name
eff: whether or not the sgRNA is included in calculation. 1: included; 0: not included
mu: mean
k: read count
loglikelihood
- [Hist of beta value]: Histogram of beta score distribution [Hist of beta value]:https://bitbucket.org/liulab/mageck_nest/downloads/Hist%20of%20beta%20value%20demo.pdf
- [QQplot of wald_p value]: The distribution of p value against uniform distribution [QQplot of wald_p value]:https://bitbucket.org/liulab/mageck_nest/downloads/QQplot%20of%20wald_p%20value%20demo.pdf
- [GSEA]: GSEA to quantify how positive control genes ranked in negatively selected hits [GSEA]:https://bitbucket.org/liulab/mageck_nest/downloads/mageck_nest_Jiyoye_PPI_major_outliers_removal_major.gene_summary_beta_1_gsea.png
