Could someone please suggest a quick way to compute the data ratio of uniquely mapped reads in
the normal to uniquely mapped reads in the tumor, as required by Varscan in the command below? I have over 50 exome BAMs.
(normal_unique_mapped_reads/tumor_unique_mapped_reads).
java -jar VarScan.jar copynumber normal-tumor.mpileup
output.basename -min-coverage 10 --data-ratio
[data_ratio] --min-segment-size 20
--max-segment-size 100
Hi, I am new and so facing problem. I was trying to make a bed graph file using bed tools genomecov command. The command was: bedtools genomecov -ibam filename.sorted.bam -g chromosome sizes.txt > O.bedgraph I got a bedgraph file which is much smaller in size. It is 500kb instead of ~6Mb. And when I load that 500kb file into IGV, I see nothing. Please help me out.
hello every one,
I have paired end illumina reads, R1.fastq and R2.fastq and I have mapped them as single-end reads by using bowtie2 default parameters, I performed further downstream analysis by using samtools and bedops, and now I have R1.bed and R2. bed I made two sets, one of them have R1_uniquely_mapped.bed, R2_uniquely_mapped.bed and second of them R1_mapped_more_than_1.bed , R2_mapped_more_than_1.bed.
because R1 and R2 belongs paired end reads, and my restriction library has maximum 2KB size, then R1 and R2 pairs must be present in less than 2 kb territory of chromosome
theoretically I am assuming, in R1.bed format,
chr1 100 180 @R1_read1______1 .................
chr1 1000 1090 @R1_read2______1................In R2.bed format,
chr1 2100 2180 @R2_read1_____2............. ## I just add 2KB length with respect to R1.bed###
chr1 2500 2590 @R2_read______2......... ## I just add 1.5KB [1500nts] with respect to R1.bed, because my library is >= 2KB.How can I customize downstream tools like BEDOPS or bedtool which can capture such type of reads or alignment????? How can I filter this type of infromation by using bedops tool????
all suggestions and comments are most welcome,
Hi,
I have a BED file with the position of retrotransposons in the mouse genome and I would like to find the nearest gene, the distance to that gene and whether it is on the + or - strand. There are so many different file formats for the mouse genome and many different databases to choose from, I was wondering what the best tool and what the best database to use would be.
Cheers, Joseph
I'd like to count the number of unique genes in a gff file falling within a list of genomic regions. With bedtools I can count the number of regions within the gff which is almost what I want, but not quite.
bedtools intersect -a regions.bed -b my.gff -cUPDATE:
I should have made my question a bit more specific. I have a modified ensemble style gtf file (not a gff) that has unique transcript IDs. This means that simply selecting unique fields in the 9th column of the gtf file actually counts transcript IDs.
To circumvent this problem I first truncated the gtf file:
cat my.gff | sed -e 's/;.*//' > delete.me.gtfThen I ran the bedtools map command:
bedtools map -a regions.bed -b delete.me.gtf -c 9 -o count_distinct > counts.genes_in_windows.bedI almost forgot to delete the intermediate file:
rm delete.me.gtfThere is probably a way to make this a oneliner, without the intermediate file, but I have a dissertation to write!
First time I am working with NGS data. I've got a BAM file with mapped reads for my samples and a BED file with the regions in HG19 that were targeted (used an Ion-torrent ampliseq panel). Are there any tools that can output something similar to this:
**Sample Amplicon Chromosome Start_coordinate_of_coverage End_coordinate_of_coverage**
Sample1 amp_001 chr6 1,000,000 1,000,250
Sample2 amp_001 chr6 1,000,111 1,000,255
Sample1 amp_002 chr6 1,000,200 1,000,333I basically want to know for each gene what coverage we have for each sample.
EDIT: changed column headings, I'm looking for coordinates that have coverage, not depth at each exon.
coverageBED -abam all.reads.bam -b refseq.genes.BED12.bed -s -split >coverage.bed
I'm confused by the mannual (Page 62):
When dealing with RNA-seq reads, for example, one typically wants to only tabulate coverage for the portions of the reads that come from exons (and ignore the interstitial intron seqeunce), The -split command allows for such coverage to be performed.If "-split" is set, the exon:exon read (for example, 30M3000N46M") exists in -abam bam file, and the 3000N will NOT be wrongly intersected when running intersectBED command. But what about coverageBED command? I do hope the 3000N will be not calculated which makes sense, and I also hope the intron body reads and other reads will be NOT ignored.Q2: If one just want to calculate exon's RPKM, does it mean one should prepare -b file to record all the exon information, and run like this:
coverageBED -abam all.reads.bam -b ...
Hi. I have a folder full of .fa files, and a .gff. The gff file contains information about which loci look like they code for RNA sequences. The .fa contain the DNA sequences for a set of human chromosomes. I want to get all the sequences which code for RNA, as defined by the gff file, out of the DNA in the fasta files. I also have a file telling me which RNA types have higher priority (lincRNA is higher priority than miRNA for example), this tells me which are more important and how I should decided between RNAs for overlapping reads in the gff.
I have been trying to code my own little program in F# that will read these files and give me each RNA read defined in the gff, and its corresponding DNA. However I am a bit confused about how it works. Do the start and end of each feature in the gff file define a character in the corresponding .fa file? Are they 1 or 0 indexed? Does it matter what strand they are ('+' or '-') for my purposes?
Ultimately my goal is to get a bunch of RNAs with their corresponding types (miRNA, lincRNA, snRNA... etc) to do some computations on.
My question is this: what is the easiest way to get it out of the data I have?
The data I am using is freely available here: http://wanglab.pcbi.upenn.edu/coral/ under the heading "Annotation packages" if anyone is interested or needs specifics.
Thank you!
I'd like to merge bed files and preserve the names of the merged features using bedtools -nms option.
However, this option (-nms) is deprecated in the newer bedtools.
The documentation says I can use -o option to get -nms behavior.
How do I get translate the new bedtools merge command to get:
bedtools merge -i file.bed -nms
I'm trying to find all the reads (by name) from a BAM file that align to various regions in a bed file. Right now I can do this with bedtools using intersectBed:
intersectBed -abam reads.bam -wo -f 1 -b regions.bed -bedFrom this one can parse all the read ids that land in every interval in regions.bed, but it's not very compact. Is there a way to get bedtools to natively transform this into a more compact format, e.g.
chr1 x y .... read_id1,read_id2,read_id3where chr1 x y is a given interval in regions.bed and the comma separated read_id1,... is the list of read ids from reads.bam that fall in that interval. In this compact format, the output BED file would have at most as many entries as there are regions in regions.bed, whereas with the -wo option it can be even larger than the number of reads in reads.bam. Thanks.
input.bam file. I think bedtools or bedops are the way to go:http://bedtools.readthedocs.org/en/latest/content/tools/bamtobed.htmlhttp://bedops.readthedocs.org/en/latest/content/reference/file-management/conversion/bam2bed.html
Other than simply running bamtobed/bam2bed, I would like to be able to define a sliding window size and step for the windows, of say, size=1000 and step=200.
I also would like to generate the bam2bed information only from a list of regions in regions.bed. E.g., something like:mapq_sliding_windows --bam input.bam --wsize 1000 -wstep 200 --regions regions.bed > mapq_sliding_windows.bed
EDITED:
Thank you Aaron for you answer. I got it working but it's slow for my 30x WGS bams:
mysql --user=genome --host=genome-mysql.cse.ucsc.edu -A -e "select chrom, size from hg19.chromInfo" > hg19.genome
bedtools makewindows -g hg19.genome -w 1000 -s 200 > hg19.windows.bed
bedtools map -a hg19.windows.bed -b <(bedtools bamtobed -i input.bam | grep -v chrM) -c 5 -o mean > ...
Hello,
I want to know if there is any way using the bedtools and miRdeep2 output bed file to get the rRNA ratio in my miRNAseq fastq data. Thank you very much!
I have a gtf file, a genome.fa, a bed file from the miRdeep2. Thanks!
Hi, I want to ask how to get the raw counts from the output of cufflinks. One way to do this is to use the fpkm.
raw counts = FPKM * (length of that transcript/1000) * (# of mapped reads / 1e6)
The FPKM and length of transcript are in the cufflinks FPKM Tracking Files. But how about the # of mapped reads?
For instance, we have a foo.bam. samtools view -c (-f|-F) flag foo.bam can do this job but I am not quite which flag should I set when it's single-end or paired-end.
Thanks!
I am using WindowBed, part of the BedTools suite, to align reads to a reference file and I obtained a very interesting result. I am trying to rule out an analysis artifact that could be caused by extending the reads or by aligning read midpoints rather than 5' ends. It is my understanding that WindowBed aligns the 5' end of the read to the reference point, rather than extending than mapping the read midpoint, or extending the 3' end of the read and mapping the midpoint. Am I correct in this assumption, that the 5' end of the read is in fact what is being aligned?
Any help here would be appreciated. The BedTools manual, which is very good, doesn't seem to address this.
Thanks
I am getting different counts for the number of bases on reference covered by aligned reads using samtools depth/mpileup and BEDTools genomeCoverageBed commands. I am using samtools-0.1.19 and bedtools-2.17.0
samtools mpileup -ABQ0 -d10000000 -f ref.fas qry.bam > qry.mpileup
samtools depth -q0 -Q0 qry.bam > qry.depth
genomeCoverageBed -ibam qry.bam -g ref.genome -dz > qry.dz
wc -l qry.[dm]*
1026779 qry.depth
1027173 qry.dz
1026779 qry.mpileupAny ideas? Thanks
Hello, I have 2 bedfiles that share some common features let's call the first file A.bed (bigger file) and the second B.bed (smaller file). I would like to have a new bed file that includes everything in B.bed in the A.bed file. I don't need the intersect, I more like need the merge option I checked bedtools's manual... couldn't find an answer for merging 2 bedfiles. Can someone help?
Thanks in advance
I have three bed files with chrNo, start, end position and type. I need to compare each chrNo, start and end position of one file with 2 other files and write the common one in a new file. Can any one suggest how can I do this efficiently? I wrote the simple perl script, but as the file is huge, it is taking a lot of time, thus is not feasible. Thanks in advance
Example files:
file1.bed:
1 20 30
1 100 120
1 200 300
file2.bed:
1 2 5
1 25 34
1 200 300
file3.bed:
1 30 33
1 200 300
1 500 600
common.bed
1 30 34 --> coordinates with overlapping 5bp is considered as same but outermost coordinates of the 3 is taken in common file
1 200 300