Uniquely mapped reads % very low

[Liyong-Zhang]

Hello Alex,

I am using STAR (STAR_2.4.2a) to mapping pair-end RNAseq data from ~100 samples, I got around 80% uniquely mapped reads for most of my samples, however, around 20 samples have extremely low values (around 0.2%). Below are my STAR mapping parameters and two log files.

for file in ${file_list[@]}
do
STAR
--runThreadN 10
--genomeDir star_index/
--readFilesCommand gunzip -c
--readFilesIn "${file}_R1_concat.fastq.gz" "${file}_R2_concat.fastq.gz"
--outFileNamePrefix "analysis/aligned_seqs/$file"
--outSAMtype BAM SortedByCoordinate
--quantMode GeneCounts
--outBAMsortingThreadN 1
done

Log file1
Started job on | May 28 12:40:29
Started mapping on | May 28 12:41:01
Finished on | May 28 12:53:45
Mapping speed, Million of reads per hour | 33.69

                      Number of input reads |       7149857
                  Average input read length |       202
                                UNIQUE READS:
               Uniquely mapped reads number |       5897884
                    Uniquely mapped reads % |       82.49%
                      Average mapped length |       200.55
                   Number of splices: Total |       4359122
        Number of splices: Annotated (sjdb) |       4086557
                   Number of splices: GT/AG |       4311072
                   Number of splices: GC/AG |       36554
                   Number of splices: AT/AC |       2054
           Number of splices: Non-canonical |       9442
                  Mismatch rate per base, % |       0.19%
                     Deletion rate per base |       0.01%
                    Deletion average length |       1.81
                    Insertion rate per base |       0.01%
                   Insertion average length |       1.40
                         MULTI-MAPPING READS:
    Number of reads mapped to multiple loci |       1056824
         % of reads mapped to multiple loci |       14.78%
    Number of reads mapped to too many loci |       2737
         % of reads mapped to too many loci |       0.04%
                              UNMAPPED READS:
   % of reads unmapped: too many mismatches |       0.00%
             % of reads unmapped: too short |       2.69%
                 % of reads unmapped: other |       0.00%

Log file2
Started job on | May 28 12:53:47
Started mapping on | May 28 12:54:23
Finished on | May 28 12:59:18
Mapping speed, Million of reads per hour | 105.82

                      Number of input reads |       8671234
                  Average input read length |       202
                                UNIQUE READS:
               Uniquely mapped reads number |       19068
                    Uniquely mapped reads % |       0.22%
                      Average mapped length |       193.63
                   Number of splices: Total |       11643
        Number of splices: Annotated (sjdb) |       10998
                   Number of splices: GT/AG |       11490
                   Number of splices: GC/AG |       100
                   Number of splices: AT/AC |       4
           Number of splices: Non-canonical |       49
                  Mismatch rate per base, % |       0.97%
                     Deletion rate per base |       0.02%
                    Deletion average length |       2.50
                    Insertion rate per base |       0.02%
                   Insertion average length |       1.94
                         MULTI-MAPPING READS:
    Number of reads mapped to multiple loci |       4579
         % of reads mapped to multiple loci |       0.05%
    Number of reads mapped to too many loci |       12
         % of reads mapped to too many loci |       0.00%
                              UNMAPPED READS:
   % of reads unmapped: too many mismatches |       0.00%
             % of reads unmapped: too short |       99.73%
                 % of reads unmapped: other |       0.00%

Thank you in advance,
Liyong