What is a HTSeq count?

The script htseq-count is a tool for RNA-Seq data analysis: Given a SAM/BAM file and a GTF or GFF file with gene models, it counts for each gene how many aligned reads overlap its exons.

How do you cite DESeq2?

Citation (from within R, enter citation(“DESeq2”) ): Love MI, Huber W, Anders S (2014). “Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2.” Genome Biology, 15, 550. doi: 10.1186/s13059-014-0550-8.

What is FPKM UQ?

Fragments Per Kilobase of transcript per Million mapped reads upper quartile (FPKM-UQ) is a RNA-Seq-based expression normalization method. The FPKM-UQ is based on a modified version of the FPKM normalization method.

What is Htseq FPKM?

Description. Fragments Per Kilobase of transcript per Million mapped reads (FPKM) is a simple expression level normalization method. The FPKM normalizes read count based on gene length and the total number of mapped reads.

What is DESeq2?

DESeq2 is a tool for differential gene expression analysis of RNA-seq data. DESeq2 is a new version of DESeq and can detect more differentially expressed genes (DEGs) than DESeq2.

How do you convert FPKM to count?

One way to convert FPKM values is to multiply the FPKM values with transcript length and the number of reads mapped in million. Trascript length can be obtained using HTSeq. I can’t understand why it is not valid to convert FPKM values into counts and use edgeR or DESeq to test for differential expression.

What is the difference between RPKM and FPKM?

The only difference between RPKM and FPKM is that FPKM takes into account that two reads can map to one fragment (and so it doesn’t count this fragment twice). TPM is very similar to RPKM and FPKM. The only difference is the order of operations.

What is normalized count?

1. The count in a list divided by the total number of observations. In the method described in this chapter, the normalized count is the score associated with a game that relates to its position in that list.

How do I run HTSeq-count from the command line?

After you have installed HTSeq (see Prequisites and installation ), you can run htseq-count from the command line: If the file htseq-count is not in your path, you can, alternatively, call the script with The are one or more files containing the aligned reads in SAM format.

What is the default read alignment name in HTSeq-count?

The default is name. If name is indicated, htseq-count expects all the alignments for the reads of a given read pair to appear in adjacent records in the input data. For pos, this is not expected; rather, read alignments whose mate alignment have not yet been seen are kept in a buffer in memory until the mate is found.

What does –nonunique None Mean in HTSeq-count?

If S is empty, the read (or read pair) is counted as no_feature. If S contains more than one feature, htseq-count behaves differently based on the –nonunique option: –nonunique none (default): the read (or read pair) is counted as ambiguous and not counted for any features.

How do the three overlap resolution modes of HTSeq-count work?

The three overlap resolution modes of htseq-count work as follows. For each position i in the read, a set S (i) is defined as the set of all features overlapping position i. Then, consider the set S, which is (with i running through all position within the read or a read pair) the union of all the sets S (i) for mode union.