title: RNA-seq of SSIIa null mutant seeds in Cadenza - rep. 3
design: Total RNA was isolated using the SpectrumTM total plant RNA kit (Sigma Aldrich, St. Louis, MO), and quanti-qualitatively assessed by Agilent 2100 Bioanalyzer RNA assay (Agilent Technologies, Santa Clara, CA). Universal Plus mRNA-Seq kit (Tecan Genomics, Redwood City, CA) was used for library preparation following the manufacturers instructions (library type: fr-secondstrand). The twelve libraries were checked with both Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) and Agilent Bioanalyzer DNA assay, and sequenced in paired-end 150 bp mode on NovaSeq 6000 (Illumina, San Diego, CA). Primary bioinformatic analysis includes base calling, demultiplexing, and adapter masking.  Raw data processing for both format conversion and de-multiplexing were carried out by Bcl2Fastq 2.20 version of the Illumina pipeline, whilst adapter sequences were masked with Cutadapt v1.11 using the following parameters: --anywhere (on both adapter sequences) --overlap 5 --times 2 --minimum- length 35 --mask-adapter).