.\" -*- mode: troff; coding: utf-8 -*- .\" Automatically generated by Pod::Man 5.01 (Pod::Simple 3.43) .\" .\" Standard preamble: .\" ======================================================================== .de Sp \" Vertical space (when we can't use .PP) .if t .sp .5v .if n .sp .. .de Vb \" Begin verbatim text .ft CW .nf .ne \\$1 .. .de Ve \" End verbatim text .ft R .fi .. .\" \*(C` and \*(C' are quotes in nroff, nothing in troff, for use with C<>. .ie n \{\ . ds C` "" . ds C' "" 'br\} .el\{\ . ds C` . ds C' 'br\} .\" .\" Escape single quotes in literal strings from groff's Unicode transform. .ie \n(.g .ds Aq \(aq .el .ds Aq ' .\" .\" If the F register is >0, we'll generate index entries on stderr for .\" titles (.TH), headers (.SH), subsections (.SS), items (.Ip), and index .\" entries marked with X<> in POD. Of course, you'll have to process the .\" output yourself in some meaningful fashion. .\" .\" Avoid warning from groff about undefined register 'F'. .de IX .. .nr rF 0 .if \n(.g .if rF .nr rF 1 .if (\n(rF:(\n(.g==0)) \{\ . if \nF \{\ . de IX . tm Index:\\$1\t\\n%\t"\\$2" .. . if !\nF==2 \{\ . nr % 0 . nr F 2 . \} . \} .\} .rr rF .\" ======================================================================== .\" .IX Title "RSEM-RUN-EBSEQ 1" .TH RSEM-RUN-EBSEQ 1 2024-03-24 "perl v5.38.2" "User Contributed Perl Documentation" .\" For nroff, turn off justification. Always turn off hyphenation; it makes .\" way too many mistakes in technical documents. .if n .ad l .nh .SH NAME rsem\-run\-ebseq \- Wrapper for EBSeq to perform differential expression analysis. .SH SYNOPSIS .IX Header "SYNOPSIS" rsem-run-ebseq [options] data_matrix_file conditions output_file .SH ARGUMENTS .IX Header "ARGUMENTS" .IP \fBdata_matrix_file\fR 4 .IX Item "data_matrix_file" This file is a m by n matrix. m is the number of genes/transcripts and n is the number of total samples. Each element in the matrix represents the expected count for a particular gene/transcript in a particular sample. Users can use 'rsem\-generate\-data\-matrix' to generate this file from expression result files. .IP \fBconditions\fR 4 .IX Item "conditions" Comma-separated list of values representing the number of replicates for each condition. For example, "3,3" means the data set contains 2 conditions and each condition has 3 replicates. "2,3,3" means the data set contains 3 conditions, with 2, 3, and 3 replicates for each condition respectively. .IP \fBoutput_file\fR 4 .IX Item "output_file" Output file name. .SH OPTIONS .IX Header "OPTIONS" .IP "\fB\-\-ngvector\fR " 4 .IX Item "--ngvector " This option provides the grouping information required by EBSeq for isoform-level differential expression analysis. The file can be generated by 'rsem\-generate\-ngvector'. Turning this option on is highly recommended for isoform-level differential expression analysis. (Default: off) .IP \fB\-h/\-\-help\fR 4 .IX Item "-h/--help" Show help information. .SH DESCRIPTION .IX Header "DESCRIPTION" This program is a wrapper over EBSeq. It performs differential expression analysis and can work on two or more conditions. All genes/transcripts and their associated statistcs are reported in one output file. This program does not control false discovery rate and call differential expressed genes/transcripts. Please use 'rsem\-control\-fdr' to control false discovery rate after this program is finished. .SH OUTPUT .IX Header "OUTPUT" .IP \fBoutput_file\fR 4 .IX Item "output_file" This file reports the calculated statistics for all genes/transcripts. It is written as a matrix with row and column names. The row names are the genes'/transcripts' names. The column names are for the reported statistics. .Sp If there are only 2 different conditions among the samples, four statistics (columns) will be reported for each gene/transcript. They are "PPEE", "PPDE", "PostFC" and "RealFC". "PPEE" is the posterior probability (estimated by EBSeq) that a gene/transcript is equally expressed. "PPDE" is the posterior probability that a gene/transcript is differentially expressed. "PostFC" is the posterior fold change (condition 1 over condition2) for a gene/transcript. It is defined as the ratio between posterior mean expression estimates of the gene/transcript for each condition. "RealFC" is the real fold change (condition 1 over condition2) for a gene/transcript. It is the ratio of the normalized within condition 1 mean count over normalized within condition 2 mean count for the gene/transcript. Fold changes are calculated using EBSeq's 'PostFC' function. The genes/transcripts are reported in descending order of their "PPDE" values. .Sp If there are more than 2 different conditions among the samples, the output format is different. For differential expression analysis with more than 2 conditions, EBSeq will enumerate all possible expression patterns (on which conditions are equally expressed and which conditions are not). Suppose there are k different patterns, the first k columns of the output file give the posterior probability of each expression pattern is true. Patterns are defined in a separate file, 'output_file.pattern'. The k+1 column gives the maximum a posteriori (MAP) expression pattern for each gene/transcript. The k+2 column gives the posterior probability that not all conditions are equally expressed (column name "PPDE"). The genes/transcripts are reported in descending order of their "PPDE" column values. For details on how EBSeq works for more than 2 conditions, please refer to EBSeq's manual. .IP \fBoutput_file.normalized_data_matrix\fR 4 .IX Item "output_file.normalized_data_matrix" This file contains the median normalized version of the input data matrix. .IP \fBoutput_file.pattern\fR 4 .IX Item "output_file.pattern" This file is only generated when there are more than 2 conditions. It defines all possible expression patterns over the conditions using a matrix with names. Each row of the matrix refers to a different expression pattern and each column gives the expression status of a different condition. Two conditions are equally expressed if and only if their statuses are the same. .IP \fBoutput_file.condmeans\fR 4 .IX Item "output_file.condmeans" This file is only generated when there are more than 2 conditions. It gives the normalized mean count value for each gene/transcript at each condition. It is formatted as a matrix with names. Each row represents a gene/transcript and each column represent a condition. The order of genes/transcripts is the same as 'output_file'. This file can be used to calculate fold changes between conditions which users are interested in. .SH EXAMPLES .IX Header "EXAMPLES" 1) We're interested in isoform-level differential expression analysis and there are two conditions. Each condition has 5 replicates. We have already collected the data matrix as 'IsoMat.txt' and generated ngvector as 'ngvector.ngvec': .PP .Vb 1 \& rsem\-run\-ebseq \-\-ngvector ngvector.ngvec IsoMat.txt 5,5 IsoMat.results .Ve .PP The results will be in 'IsoMat.results' and 'IsoMat.results.normalized_data_matrix' contains the normalized data matrix. .PP 2) We're interested in gene-level analysis and there are 3 conditions. The first condition has 3 replicates and the other two has 4 replicates each. The data matrix is named as 'GeneMat.txt': .PP .Vb 1 \& rsem\-run\-ebseq GeneMat.txt 3,4,4 GeneMat.results .Ve .PP Four files, 'GeneMat.results', 'GeneMat.results.normalized_data_matrix', 'GeneMat.results.pattern', and 'GeneMat.results.condmeans', will be generated.