HOMER

homerTools [command] [command specific options]

i.e. homerTools trim -3 AAAAAAAA s_1_sequence.txt

barcodes - for separating and removing 5' barcodes from FASTQ/FASTA files
trim - for trimming by adapter sequence, specific lengths, etc. from FASTQ/FASTA files
freq - for calculating nucleotide frequencies in FASTQ/FASTA/txt sequence files
extract - for extracting specific regions of seqeuence from genomic FASTA files

To separate and remove 5' barcodes from sequencing data (where the first "x" base pairs of the read are the barcode):



The 3rd argument must be the length of the 5' barcode, which will be the first base pairs in the sequence.  By default, this command creates files named "filename.barcode", such as s_1_sequence.txt.AAA, s_1_sequence.txt.AAC, s_1_sequence.txt.AAG etc.  The parameter "-min <#>" specifies the minimum barcode frequence to keep (default is 0.02 [2%]).  The frequency of each barcode is recorded in the output file "filename.freq.txt".  If important barcodes were deleted, rerun the command with a smaller value for "-min <#>".

With all the fancy types of sequencing being done, it is getting common to find adapters as part of the sequences that are analyzed.  The trim command allows users to trim sequences from the 3' and 5' ends by either a specific number of nucleotides or remove a specific adapter sequence.  The basic command is executed like this:


The output will be placed in files "filename.trimmed" and the distribution of sequence lengths after trimming will be in "filename.lengths" for each of the input files.  The following options control how homerTools trims the sequences:


For adapter sequence trimming, it will search for the first full match to the sequence and delete the rest of the sequence.  For example if you specify "-3 AA", it will search for the first instance of "AA" and delete everything after it.  It will also delete partial matches if they are at the end of the sequence (or beginning for 5').  As another example, our lab uses an amplification strategy for RNA that results in the ligation of a polyA tail to the RNA sequence.  If the reads are long enough, the read will be just As.

The extract command can be used to extract large numbers of specific genomic sequence.  The first input file you need is a HOMER style peak file or a BED file with genomic locations.  Next, you must have the genomic DNA sequences in one of two formats: (1) a directory of chr1.fa, chr2.fa FASTA files (can be masked file like *.fa.masked), or (2) a single file FASTA file with all of the chromosomes concatonated in one file.  The sequences are sent to stdout as a tab-delimited file, or as a FASTA formatted file if "-fa" is added to the end of the command.  Save the output to a file by adding " > outputfile.txt" to the end of the command. The program is run like this:

The freq command will calculate nucleotide frequencies from FASTQ, FASTA, or tab-delimited text sequence files.  The program tries to auto detect the format, but it may help to specify the format directly ("-format fastq", "-format fasta", "-format tsv").  The program outputs a position-dependent nucleotide/dinucleotide frequency file as a function of the distance from the start of the sequencing reads.  The output is sent to stdout, unless you specify "-o <outputfile.txt>".  If you specify "-gc <outpufile2.txt>", the program will also create a file that specifies the cumulative frequency of CpG, total G+C,  total A+G, and total A+C in each individual sequence.