I am mapping rna-seq reads to a reference genome. Very sure, the reads and the genome reference are generated from same variety. They are both downloaded from internet. I firstly trimmed the paired reads using trimmomatic, by cutting the TruSeq_adapters, cutting the LEADING and TRAILING bases with quality score lower than 4, and removing the low quality score reads (SLIDINGWINDOW:4:15). Then I checked the reads using fastqc. The filtered reads looks fine. I am sure the adapters removed and average quality score per read is 37, and no read with score lower than 26.
Then I run the STAR with default parameter. Result summary: 9472216 reads; of these: 9472216 (100.00%) were paired; of these: 6353973 (67.08%) aligned concordantly 0 times 2570774 (27.14%) aligned concordantly exactly 1 time 547469 (5.78%) aligned concordantly >1 times ---- 6353973 pairs aligned concordantly 0 times; of these: 8552 (0.13%) aligned discordantly 1 time ---- 6345421 pairs aligned 0 times concordantly or discordantly; of these: 12690842 mates make up the pairs; of these: 12622901 (99.46%) aligned 0 times 51272 (0.40%) aligned exactly 1 time 16669 (0.13%) aligned >1 times 33.37% overall alignment rate
Only (27.14%) aligned concordantly exactly 1 time.
I also tried other methods such as Hisat2. It shows ".....2570774 (27.14%) aligned concordantly exactly 1 time......." .
I don't know whether this result is common. But I think the percentage of Uniquely mapped reads may be too low. The data are from an allopolyploid plant. Any comment a good help to me.