I have problem in the trimming my RNA seq reads. The problem is that although the length of all reads are the same, 101 nucleotids, quality test by fastqc shows some reads contain adapter (adapter content is red). For reads containing adapters, I trimmed them by Trimmomatic using ILLUMINACLIP command resulting in the adapter removal. My problem is for some other reads that I can not understand if they contain adapter or not because the adapter content in fastqc file is yellow and according to adapter content graph, from 12th or 13th nucleotide, it seems something wrong because before 12th nucleotide, the graph line is completely direct. I trimmed the reads according to mRNA-Seq Library Prep Kit V2 Lexogen protocol as "The first nine nucleotides need to be removed from Read 1 (starter side), while on the stopper side it is only six nucleotides (Read 2)." But fastqc test shows the same result as before and the problem still exists. So I got completely confused. Would you pleas help me to trim these reads?
You appear to be confusing trimming Illumina adapters and the specific trimming that Lexogen has recommended. Instructions from lexogen appear to remove specific adapters/nucleotides their kit must be adding to the fragments. You must not have trimmed your reads correctly with Trimmomatic to remove the Illumina adapter. Based on that graph your reads after proper trimming will have a range of lengths (i.e. they won't remain all 101 bp).
Lexogen's mRNA-Seq protocol uses random-primer. AFAIK, they suggest to remove them, if you want to use Tophat2 or something similar sensitive. If you use STAR or BBmap, you can keep the complete read but you should increase the allowed numbers of mismatches.
FastQC needs a certain length to identify adapter sequences. This is apparently 12 or 13 nts. Regardless of how you trim the reads' start, the detection of adapter sequences will start at this position. Denote, the graph of the adapter content also ends before the actual cycle number is reached.
You can use fastp to preprocess your Illumina sequencing data (no matter RNASeq / DNASeq, no matter PE/SE). It can trim adapters automatically for both PE and SE data, which means that you don't have to input the adapter sequences.
Besides trimming adapters, this tool also performs quality filtering and other operations to improve your data quality. And most of the features are automated. All you have to do is to install fastp, and run: