有网友咨询过对于没有参考基因组或者转录组的物种，如何做RNA-seq分析。我觉得这个问题太大了，而且我还真的对这个没有经验。但是我以前看到过一篇文献，里面提到过一个非常全面的转录组 de novo组装注释流程，所以我摘抄了文章里面的生物信息学处理部分，分享给大家：
Raw reads from all three data sets were filtered to remove adapter sequences with sequence pre-processing tool, Trimmomatic . High quality Illumina raw reads with phred score ≥ 25 were kept for assembly. De novo assembly of these processed reads was performed with Trinity (v2.0.6) . Statistics of the assembly is showed in Table 1.
- Trinotate http://trinotate.github.io download Trinotate
- Trinity (includes support for expression and DE analysis using RSEM and Bioconductor): http://trinityrnaseq.github.io/ download Trinity. >Note, Trinity is not absolutely required. It is possible to use Trinotate with other sources of transcript data as long as suitable inputs are available.
- TransDecoder for predicting coding regions in transcripts http://transdecoder.github.io download TransDecoder.
- sqlite (required for database integration): http://www.sqlite.org/
- NCBI BLAST+: Blast database Homology Search: http://www.ncbi.nlm.nih.gov/books/NBK52640/
- HMMER/PFAM Protein Domain Identification: http://hmmer.janelia.org/download.html