Puzzles to help teach you more bedtools.

  1. Create a BED file representing all of the intervals in the genome that are NOT exonic.

Answer:

bedtools complement -i exons.bed -g genome.txt > notexons.bed
  1. What is the average distance from GWAS SNPs to the closest exon? (Hint - have a look at the closest tool.)

Answer:

bedtools closest -a gwas.bed -b exons.bed -d | head
chr1    1005805 1005806 rs3934834   chr1    1007125 1007955 NM_001205252_exon_0_0_chr1_1007126_r    0   -   1320
chr1    1079197 1079198 rs11260603  chr1    1078118 1079434 NR_038869_exon_2_0_chr1_1078119_f   0   +   0
chr1    1247493 1247494 rs12103 chr1    1247397 1247527 NM_001256456_exon_1_0_chr1_1247398_r    0   -   0
chr1    1247493 1247494 rs12103 chr1    1247397 1247527 NM_001256460_exon_1_0_chr1_1247398_r    0   -   0
chr1    1247493 1247494 rs12103 chr1    1247397 1247527 NM_001256462_exon_1_0_chr1_1247398_r    0   -   0
chr1    1247493 1247494 rs12103 chr1    1247397 1247527 NM_001256463_exon_1_0_chr1_1247398_r    0   -   0
chr1    1247493 1247494 rs12103 chr1    1247397 1247527 NM_017871_exon_1_0_chr1_1247398_r   0   -   0
chr1    2069171 2069172 rs425277    chr1    2066700 2066786 NM_001033581_exon_1_0_chr1_2066701_f    0   +   2386
chr1    2069171 2069172 rs425277    chr1    2066700 2066786 NM_001033582_exon_1_0_chr1_2066701_f    0   +   2386
chr1    2069171 2069172 rs425277    chr1    2066700 2066786 NM_001242874_exon_1_0_chr1_2066701_f    0   +   2386

bedtools closest -a gwas.bed -b exons.bed -d \
  | awk '{ sum += $11 } END { if (NR > 0) print sum / NR }'
46713.1
  1. Count how many exons occur in each 500kb interval (“window”) in the human genome. (Hint - have a look at the makewindows tool.)

Answer:

bedtools makewindows -g genome.txt -w 500000 > genome.windows.bed
bedtools intersect -a genome.windows.bed -b exons.bed -c > genome.windows.exoncount.bedg

or…

bedtools makewindows -g genome.txt -w 500000 \
  | bedtools intersect -a - -b exons.bed -c \
> genome.windows.exoncount.bedg
  1. Are there any exons that are completely overlapped by an enhancer? If so, how many?

Answer:

bedtools intersect -a exons.bed \
                   -b <(grep Enhancer hesc.chromHmm.bed) \
                   -wa -wb -f 1.0 \
| head
chr1    948846  948956  NM_005101_exon_0_0_chr1_948847_f    0   +   chr1    948337  949337  4_Strong_Enhancer
chr1    1051439 1051736 NM_017891_exon_9_0_chr1_1051440_r   0   -   chr1    1051337 1051737 6_Weak_Enhancer
chr1    1109285 1109306 NM_001130045_exon_0_0_chr1_1109286_f    0   +   chr1    1108537 1109537 6_Weak_Enhancer
chr1    1109803 1109869 NM_001130045_exon_2_0_chr1_1109804_f    0   +   chr1    1109737 1109937 6_Weak_Enhancer
chr1    1219357 1219470 NM_001130413_exon_4_0_chr1_1219358_f    0   +   chr1    1219137 1220137 7_Weak_Enhancer
chr1    1219357 1219470 NR_037668_exon_4_0_chr1_1219358_f   0   +   chr1    1219137 1220137 7_Weak_Enhancer
chr1    1229202 1229313 NM_030649_exon_1_0_chr1_1229203_r   0   -   chr1    1228937 1229937 6_Weak_Enhancer
chr1    1229469 1229579 NM_030649_exon_2_0_chr1_1229470_r   0   -   chr1    1228937 1229937 6_Weak_Enhancer
chr1    1234724 1234736 NM_030649_exon_14_0_chr1_1234725_r  0   -   chr1    1234137 1234937 7_Weak_Enhancer
chr1    1245060 1245231 NM_153339_exon_4_0_chr1_1245061_f   0   +   chr1    1244937 1245337 4_Strong_Enhancer

bedtools intersect -a exons.bed \
                   -b <(grep Enhancer hesc.chromHmm.bed) \
                   -wa -wb -f 1.0 -u \
| wc -l
13746
  1. What fraction of the GWAS SNPs are exonic?

Answer:

wc -l gwas.bed
17680 gwas.bed

bedtools intersect -a gwas.bed -b exons.bed -u | wc -l
1625

echo "foo" | awk '{print 1625/17680}'
0.0919118
  1. What fraction of the GWAS SNPs are lie in either enhancers or promoters in the hESC data we have?

Answer:

bedtools intersect -a gwas.bed -b <(egrep "Enhancer|Promoter" hesc.chromHmm.bed) -u \
| wc -l
1285

echo "foo" | awk '{print 1285/17680}'
0.072681
  1. Create intervals representing the canonical 2bp splice sites on either side of each exon (don’t worry about excluding splice sites at the first or last exon). (Hint - have a look at the flank tool.)

Answer:

bedtools flank -l 2 -r 2 -i exons.bed -g genome.txt > splice-sites.bed

Or:

bedtools slop -b 2 -i exons.bed -g genome.txt > exons.plus2.bed
bedtools subtract -a exons.plus2.bed -b exons.bed > splice-sites.bed
  1. What is the Jaccard statistic between CpG and hESC enhancers? Compare that to the Jaccard statistic between CpG and hESC promoters. Does the result make sense? (Hint - you will need grep).

Answer:

bedtools jaccard -a cpg.bed -b <(grep Enhancer hesc.chromHmm.bed)
intersection    union-intersection  jaccard n_intersections
1148180 132977386   0.0086344   4969

bedtools jaccard -a cpg.bed -b <(grep Promoter hesc.chromHmm.bed)
intersection    union-intersection  jaccard n_intersections
15661111    53551816    0.292448    20402
  1. What would you expect the Jaccard statistic to look like if promoters were randomly distributed throughout the genome? (Hint - you will need the shuffle tool.)

Answer:

bedtools shuffle -i <(grep Promoter hesc.chromHmm.bed) -g genome.txt \
  | sort -k1,1 -k2,2n \
> promoters.shuffled.bed

bedtools jaccard -a cpg.bed -b promoters.shuffled.bed
intersection    union-intersection  jaccard n_intersections
294071  68556207    0.00428949  78
  1. Which hESC ChromHMM state (e.g., 11_Weak_Txn, 10_Txn_Elongation) represents the most number of base pairs in the genome? (Hint: you will need to use awk or perl here, as well as the groupby tool.)