RUFUS k-mer based genomic variant detection
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K-mer based variant detection.

Andrew Farrell PhD
Research Associate
Department of Human Genetics
USTAR Center for Genetic Discovery
Eccles Institute of Human Genetics
University of Utah School of Medicine​ 15 North 2030 East, Room 7140
Salt Lake City, UT 84112-5330

This project is still under development and its not considered stable but you are free to use and any feedback is welcome.

Compiler Requirements

RUFUS requires the use of the gcc/4.9.2 compiler. If you are not using the gcc/4.9.2 compiler, RUFUS will not build and install properly. Please make sure that are using the gcc/4.9.2 compiler before you proceeed.

Download and install

git clone   

This should install everything you need to use RUFUS. If you get errors during the installation contact me at at or submit an issue. NOTE, to date this only works on linux machines, we have not optimized for other platforms

Testing RUFUS

To make sure that RUFUS was successfully built, we provide users with a test run script to run RUFUS on a small test set of data with a small test reference, and default parameters. To test RUFUS, simply run

cd testRun

Make sure that is called directly from the testRun directory, or the testRun script will not be able to find the appropriate resources.

All data for this run is contained in the resources dir, and nothing needs to be provided by the user to test RUFUS.

At the end of a successfull test run, you should see a file named


This file should contain a single varient call. The call should look exactly as follows:

#CHROM  POS     ID      REF     ALT     QUAL    FILTER  INFO    FORMAT  ./Child.bam     Mother.bam      Father.bam
5:177630000     12896   X-DeNovo        T       G       25      PASS    RN=NODE_Child.bam.generator.V2_0_L273_D22:10:12::MH0;MQ=60;cigar=273M;SB=0.454545;CVT=X;HD=-1_-1_-1_-1_-1_19_-1_19_19_-1_-1_-1_-1_-1_20_20_19_-1_-1_18_-1_18_-1_-1_18_-1_-1_;AO=19;VT=X GT:DP:RO:AO     0/1:39:20:19    0/0:23:23:0     0/0:23:23:0

If you were unable to reproduce this call, something went wront with the RUFUS install, and you should get the test run to work before proceeding further. If you are unable to reproduce a successfull test run, please contact me at JAndrewRFarrell@gmail.scom


RUFUS is primarily used to find mutations unique to a proband sample, that are not found in the control samples


./ [-s|--subject <arg>] [-c|--controls][<controls-1>] ... [<controls-n>]  [-t|--threads <arg>] [-k|--kmersize <arg>] [-r|--ref <arg>] [-m|--min <arg>] 
-s,--subject: bam file containing the subject of interest (REQUIRED)

-c, --controls: bam files containing the control subjects (REQUIRED)

-t,--threads: number of threads to use (REQUIRED)

-k,--kmersize: size of Kmer to use (REQUIRED)

-r,--ref: file path to the desired reference file to create VCF (REQUIRED)

-m,--min: overwrites the minimum k-mer count to call variant (OPTIONAL, Do not provide a min unless you are sure what you want)

-h,--help: HELP!!!!!!!!!!!!!!!

The command line should look something like this:

bash --subject Child.bam --controls Mother.bam Father.bam  --kmersize 25 --threads 40 --ref human_reference_v37_decoys.fa


bash -s Child.bam -c Mother.bam Father.bam -k 25 -t 40 -r human_reference_v37_decoys.fa

The flags can be provided any any order.

RUFUS can take any number control files (Must provide atleast one).

Provide all of your control files after the [-c|--controls] flag

For Example:

bash -s tumorT1.bam -c tumorT0.bam -k 25 -t 40 -r human_reference_v37_decoys.fa


bash -s Proband.bam -c Mother.bam Father.bam Sibling1.bam Sibling2.bam -k 25 -t 40 -r human_reference_v37_decoys.fa

We recommend a kmer size of 25, 40 threads, and to NOT provide RUFUS with the optional --min parameter

Providing a reference file.

After RUFUS has identified reads containing mutant kmers, the reads must be aligned to a reference fasta file. Any fasta file can be used as a reference, as long as the fasta file has been indexed for BWA. If a fasta has been indexed by bwa, there will be reference files with the following extensions: pac, .ann, .abm, .bwt, sa. In order to prepare a reference fasta for bwa, simply type:

bwa index -a bwtsw reference.fa
samtools faidx reference.fa

This will produce the BWA index files, and the fasta file index respectively. Make sure that the bwa index files and the fasta index file are in the same directory as reference.fa