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Pipeline publication

[…] he repeat masked genome data based on Tophat2 with default options (). The aligned results with transcript data were implicated in prior gene prediction by using GeneMark-ET tool (v4.29; ). Next, homologous proteins of other species were mapped to the genome using TBlastN (Blast v2.2.29) with an E-value cutoff of 1E–5 and the aligned protein sequences were implicated in prediction of the gene region using Exonerate tool v2.2.0 with default parameters (; ). A final gene set of R. philippinarum was decided by using AUGUTUS tool v3.2.1 with default setting (). Gene functions were assigned according to the best alignment attained using BLASTP to the UniProt database (E-value cutoff of 1E–5) and INTERPRO v5.17 scan., We collected C1qDC genes from the R. philippinarum and C. gigas genomes, and used the Pfam seed alignment sequences of C1q gene family (PF00386) as their conserved sequences (). Next, the ClustalW2 multiple alignment program was used to generate the multiple amino acid sequence alignment of the C1q domain gene using default option (). Through the Jalview v2.10.1 program, the conserved domains of the C1q gene were selected based on the Pfam seed alignment sequences and the phylogenetic tree was constructed with the BLOSUM62 and the Neighbor-Joining methods (). The generated tree was finally visualized by using FigTree tool v1.4.3 (, In order to construct cDNA library for RNA sequencing, mRNA was enriched by oligo-dT attached magnetic beads from a total 2 µg of RNA. Then, the purified mRNAs were sheared into short fragments and synthesized to double-stranded cDNA by reverse-transcription, immediately. Synthesized cDNA was subjected to end-repair, poly-A addition, and connected with 5′ and 3′ adaptors on both ends using the TruSeq RNA sample prep Kit (Illumina, CA). Modified mRNA fragments were separated on bluepippin 2% agarose gel cassette and the suitable fragments were automatically purified and used as templates for PCR amplification. The final produc […]

Pipeline specifications

Software tools InterPro, Clustal W, Jalview, FigTree