Rubus GDR RefTrans V1
Materials & Methods
GDR Rubus RefTrans combines published RNA-Seq and EST data sets to create a reference transcriptome (RefTrans) for rubus and provides putative gene function identified by homology to known proteins.
In Rubus_RefTrans_V1, 265 million Illumina paired-end reads from publicly available peer-reviewed rubus RNA-Seq data sets (Hyun et al. 2014, Hyun et al. 2014, Garcia-Seco et al. 2015), and 3,184 ESTs, were downloaded from the NCBI Short Read Archive database (SRR975478, SRX347804), the EBI database (PRJEB6680) and the NCBI dbEST database, respectively. The RNA-Seq sequences were subjected to quality control using the NGS QC Toolkit (V2.3.3, default parameters, Patel and Jain, 2012) and custom Perl scripts. The remaining 264 million RNA-Seq reads were assembled de novo with Trinity (v2.0.6, Grabherr et al, 2011) using default assembly parameters and a minimum coding length of 200 bases. Quality control of the ESTs included vector sequence screening (UniVec_Core,ftp://ftp.ncbi.nih.gov/pub/UniVec/) using cross_match (Gordon et al, 1998), removal of tRNA/rRNA/snRNA sequences identified using tblastx (Altschul et al, 1990), and Poly-A tail trimmimg. The filtered ESTs were assembled using the CAP3 program (P -90, Huan and Madan, 1999). Bowtie (v 2-2.2.3) (Langmead et al, 2009) was applied to multi-map the RNA-Seq reads and ESTs back to the assembled contigs and singlets. The contigs and singlets were hierarchically clustered into genes using Corset (v1.0.4) (Davidson and Oshlack, 2014) with default parameters. The longest isoform was selected to represent each Corset cluster, creating a RefTrans V1 for rubus of 37,326 sequences. The RefTrans were functionally characterized by pairwise comparison using the BLASTX algorithm against the Swiss-Prot (UniProtKB/Swiss-Prot Release 2015_10) and TrEMBL (UniProtKB/TrEMBL Release 2015_10) (Boeckmann et al, 2003) protein databases. Information on the top 25 matches with an expect (E) value of ≤ 1E-06 were recorded and stored in the database. The transcriptome and annotation (GO Terms, match description, InterPro domains) are available for searching and downloading.