To many clusters without any count values

[Sktbanerjee1]

I have recently started using LeafCutter for sQTL analysis. I have used regtools to generate the junction files with the following command regtools junctions extract -a 8 -m 50 -M 500000 -s 1 $bamfile -o $bamfile.junc. The list of these junction files are then being used to generate the clusters using the following code:

python ${leafcutter_dir}/clustering/leafcutter_cluster_regtools.py \
-j ${junc_files} \
-m 50 \
-o intron_clusters \
-r ${out_dir} \
-l 500000

The cluster counts that I get look strange as for some clusters the values are continuously zero across all of the samples. Additionally, the cluster counts seem to be varying drastically across the samples.

Here is a screenshot of some of the rows

1:4492668:4492840:clu_1_- 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 6 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 0 0 0 0 4 0 0 0 0 0 3 0 0
 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 2 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 7 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 0 0 0 
2 0 1 0 0 0 0 0 0 0 0 0 0 4 1 0 0 0 0 0 0 0 0 0 0 1 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 0 0 6 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
 0 0 0 0 0 0 0 1 3 0 0 0 0 0 0 0 0 2 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 2 2 0 0 0 0 0 
0 1 0
1:4492668:4493100:clu_1_- 4 29 0 18 22 22 5 0 0 18 0 26 12 0 10 27 0 0 0 12 25 6 10 20 16 8 34 25 17 13 0 10 26 21 0 0 16 13 9 18 15 21 0 10 12 11 13 12 42 12 66 7 12 22 0 28 0 36 11 0 9 14 11 22
 11 11 5 25 0 21 4 10 0 25 0 13 16 28 26 0 20 6 25 9 0 0 29 23 0 0 18 7 5 16 21 11 14 0 11 12 12 32 0 32 23 0 18 0 13 0 0 3 22 25 0 17 24 4 0 0 18 5 0 13 19 17 0 12 0 28 11 15 34 8 0 0 17 36 16 1
2 0 16 5 0 0 9 24 20 3 21 0 28 21 20 22 29 6 14 35 13 10 0 0 0 9 0 8 0 0 0 13 21 24 0 7 23 12 20 26 0 15 0 22 6 16 16 10 5 12 12 13 0 18 0 0 19 18 0 15 16 37 15 6 0 0 9 12 6 0 22 15 21 17 5 0 8 0
 12 0 16 47 0 23 35 12 14 9 5 0 31 12 5 0 4 0 9 0 40 13 12 9 34 9 13 30 42 8 14 14 0 9 0 8 16 2 13 8 15 54 0 12 10 29 10 0 0 5 17 19 14 0 0 2 14 14 0 14 17 12 0 22 0 15 10 0 15 0 9 21 62 7 30 0 1
2 24 0 47 37 7 13 0 0 17 0 0 22 14 23 17 13 9 5 6 27 8 0 0 9 50 0 34 18 13 7 14 11 13 6 10 17 0 23 0 11 23 2 25 23 0 0 17 31 39 23 17 28 7 34 20 0 4 0 2 18 7 7 11 16 0 27 32 9 6 15 23 10 8 6 4 11
 7 36 6 30 7 29 15 10 2 7
1:4492668:4493772:clu_1_- 0 1 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 6 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 0 0
 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 4 2 0 0 0 0 0 0 0 0 0 0 0 6 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 
0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 2 0 1 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 5 0 0 0 0 0 0 3 0 0 0
 0 0 0 0 0 0 0 0 0 1 0 3 0 0 0 0 1 0 0 0 0 0 0 0 0 9 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 2 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 2 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 
0 0 0
1:4493466:4493772:clu_1_- 0 0 0 0 5 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 2 1 0 2 4 0 0 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 3 0 0 0 0 0 0 2 0 0 2 1 0 0 1 1 0 0 0 0 0 0 0 1 0 1 0 0 0 0 0 1 0 0 0
 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 2 0 0 0 0 1 0 0 0 0 0 1 0 0 1 0 1 0 0 4 0 0 0 0 0 0 0 0 2 2 0 0 0 0 0 0 1 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 5 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 2 0 0 0 0 5 0 
0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 4 0 0 0 3 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 8 1 8 0 0 0 0 0 0 0 1 0 1 0 0 3 0 0 0 1 1 0 0 1 1 0 0 0 0 0 0 0 0 3 1 0 0
 0 0 0 0 0 1 0 0 0 3 2 1 0 0 4 0 3 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 3 0 1 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 4 0 0 0 0 6 2 0 0 0 2 8 0 0 0 0 3 0 0 1 0 0 0 5 0 0 0 5 0 1 1 0 3 0 0 0 0 0 0 0 
0 0 0
1:4493466:4495136:clu_1_- 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 0 1 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 4 2 0 2 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0
 0 2 0 0 0 2 0 0 0 0 2 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 4 3 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 
1 0 2 0 0 2 1 0 1 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 1 0 1 0 1 0 1 0 2 0 2 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 2 0 0 0 0 0 0 0 0 1 0
 0 0 0 0 0 1 0 1 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 0 1 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 2 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 2 0 0 0 0 1 
0 0 0
1:4493490:4493772:clu_1_- 0 2 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 2 0 0 1 0 0 3 6 0 0 0 0 0 0 0 0 0 1 5 0 0 0 0 0 0 2 0 0 0 0 2 0 0 0 0 0 0 0 0 0 2 0 0 2 0 0 0 0 0 2 0 0 0 4 0 2 1 0 0 0 0 2 1 0 0
 0 0 0 0 0 0 0 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 4 1 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 3 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 
0 0 1 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 4 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 0 0 1 0 0 4 0 0 1 0 0 0 0 0 0 0 0 1 0 0 1 0 0 11 0 0 2 1 0 0 0 0 1 0 0 1 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 
0 3 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 5 0 0 0 0 1 1 0 0 0 0 0 0 0 1 0 0 0 0 2 0 0 0 0 0 0 3 0 4 0 3 0 0 0 0 1 0 2 0 0 0 0 0 0 0 0 1 6 0 0 1 0 0 0 0 0 0 0 0 0
 0 0 0
1:4493863:4495136:clu_1_- 2 3 0 11 9 6 9 0 0 9 0 1 5 0 6 0 0 0 0 5 6 5 2 9 7 3 8 15 3 2 0 3 5 3 0 0 8 1 1 3 1 6 0 3 3 4 3 0 6 14 4 10 4 2 0 3 0 15 0 0 3 2 0 0 7 4 0 6 0 1 0 0 0 11 0 14 2 6 8 0 3 
3 4 2 0 0 0 15 0 0 0 1 2 7 3 0 1 0 4 6 0 5 0 2 10 0 1 0 1 0 0 5 17 2 0 1 0 4 0 0 9 2 0 8 13 2 0 0 0 3 5 0 11 0 0 0 2 5 5 2 0 0 3 0 0 0 3 1 1 2 0 0 5 2 17 1 7 1 6 7 1 0 0 0 3 0 2 0 0 0 2 7 0 0 1 4
 2 4 3 0 9 0 6 9 5 0 0 0 2 18 1 0 5 0 0 0 3 0 11 5 0 0 4 0 0 4 0 0 0 12 4 8 2 0 0 3 0 3 0 2 12 0 0 2 8 6 8 6 0 0 1 0 0 1 0 0 0 3 6 0 2 4 9 2 10 8 0 0 1 0 2 0 1 4 0 2 3 4 6 6 3 6 4 0 0 0 1 10 0 6 
0 0 0 4 2 0 3 3 0 0 16 0 0 1 0 3 0 3 0 6 0 5 0 1 5 0 5 7 1 7 0 0 1 0 0 12 1 10 3 0 0 1 4 2 0 0 0 8 10 0 9 3 4 4 5 4 3 0 3 2 0 2 0 4 6 2 11 6 0 0 1 4 14 8 3 7 0 11 12 0 4 0 5 6 12 4 7 3 0 0 16 4 0
 1 0 4 1 4 1 8 1 10 0 0 6 2 1 1 0 0
1:4493863:4496291:clu_1_- 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0
 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 
0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 2 3 0 0 0 0 0 0 0 0 4 0 0 0 0 0 0 0 0 0 0 4 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 2 6 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0
 0 0 0 0 0 0 0 1 0 0 0 0 0 0 5 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 1 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 3 0 0 0 0 0 0 1 0 0 1 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 1 2 1 0 0 0 0 0 0 0 0 0 0 1 
0 0 0
1:4495198:4496291:clu_1_- 1 2 0 4 0 4 7 0 0 1 0 3 3 0 0 0 0 0 0 0 0 3 3 6 3 4 6 10 1 0 0 0 9 2 0 0 4 1 4 1 2 3 0 0 0 3 2 6 6 1 0 11 1 4 0 0 0 14 2 0 5 6 2 5 0 1 0 3 0 14 2 3 0 3 0 3 2 8 10 0 2 0 
3 0 0 0 2 14 0 0 2 0 1 5 5 3 2 0 2 3 1 2 0 7 5 0 0 0 1 0 0 1 3 0 0 1 2 4 0 0 0 0 0 7 17 1 0 0 0 6 3 1 3 1 0 0 0 6 4 5 0 1 4 0 0 3 3 3 1 2 0 0 1 0 10 5 6 0 6 4 2 0 0 0 1 0 0 0 0 0 0 1 8 0 0 6 0 4 
6 0 3 0 3 10 6 1 0 1 1 11 5 0 10 0 0 11 1 0 3 3 2 2 2 0 0 3 0 1 0 0 2 5 0 0 0 2 0 6 0 4 11 0 5 8 5 3 4 3 0 3 2 1 0 3 0 2 0 4 3 3 3 3 10 2 6 0 0 1 4 0 0 0 3 3 0 0 4 5 13 0 9 1 7 0 0 0 0 5 0 8 0 0 
0 1 4 0 0 6 1 0 10 0 1 2 0 8 0 3 0 7 1 4 0 1 0 0 8 1 0 5 0 0 1 0 0 5 0 1 1 0 0 2 3 7 2 0 0 2 0 0 2 4 0 4 1 0 5 1 5 5 0 1 0 10 4 1 0 0 0 0 4 8 4 0 3 1 2 4 9 0 3 0 4 1 3 2 5 2 0 4 6 2 1 5 1 0 2 0 2
 0 2 10 1 7 0 2 4 0 0 0

I have no experience with Leafcutter, so just wondering If this is normal.

Also, I looked into the BAM files within IGV and some of the clusters, where the count is zero, I can still see junction reads in IGV. I am wondering, how is that possible. I will be grateful if you can kindly help me understand what's going on.