Abstract
Uniquely, oat, among cereals, accumulates an appreciable amount of oil in the endosperm together with starch. Oat is also recognized for its soluble fibers in the form of β-glucans. Despite high and increasing interest in oat yield and quality, the genetic and molecular understanding of oat grain development is still very limited. Transcription factors (TFs) are important regulatory components for plant development, product quality and yield. This study aimed to develop a workflow to determine seed tissue specificity of transcripts encoding transcription factors to reveal differential expression of potential importance for storage compound deposition and quality characters in oat. We created a workflow through the de novo assembly of sequenced seed endosperm and embryo, and publicly available oat seed RNAseq dataset, later followed by TF identification. RNAseq data were assembled into 33,878 transcripts with approximately 90% completeness. A total of 3875 putative TF encoding transcripts were identified from the oat hybrid assemblies. Members of the B3, bHLH, bZIP, C3H, ERF, NAC, MYB and WRKY families were the most abundant TF transcripts. A total of 514 transcripts which were differentially expressed between embryo and endosperm were identified with a threshold of 16-fold expression difference. Among those, 36 TF transcript homologs, belonging to 7 TF families, could be identified through similarity search in wheat embryo and endosperm EST libraries of NCBI Unigene database, and almost all the closest homologs were specifically expressed in seed when explored in WheatExp database. We verified our findings by cloning, sequencing and finally confirming differential expression of two TF encoding transcripts in oat seed embryo and endosperm. The developed workflow for identifying tissue-specific transcription factors allows further functional characterization of specific genes to increase our understanding of grain filling and quality.
Highlights
Oat (Avena sativa) is a cereal grown for its grain and is cultivated in Europe, North America, Russia, Australia and northern China
Libraries were prepared at BGI (Shenzhen, China) using oligo(dT)-enriched mRNA subsequently fragmented and first-strand cDNA synthesis by random hexamer–primer according to their library preparation protocol and oat embryo and endosperm RNA libraries were sequenced through Illumina sequencing platform HiSeq 2000 as unpaired-end reads
Embryo and endosperm-specific transcriptome was generated from newly sequenced endosperm and embryo reads (SE) through Trinity and Bridger de novo transcriptome assembler and assemblies generated using both assemblers were clustered through CD-HIT clustering software to generate a non-redundant set of transcripts
Summary
Oat (Avena sativa) is a cereal grown for its grain and is cultivated in Europe, North America, Russia, Australia and northern China. Oat is well known as an excellent source of β-glucan (1–3; 1–4 mixed link β-d-glucan), a dietary fiber with documented and approved health claims regarding blood glucose stabilizing and cholesterol lowering properties (Butt et al 2008; Rasane et al 2015; Chen et al 2016). These different compounds are accumulating in the oat grain in which the endosperm tissue makes out the major and the embryo tissue only a minor part. Even though great progress has been made in bioinformatics data analysis, molecular data of non-model plants are still imposing new challenges to methods of bioinformatics data analysis and approaches for the genetic and molecular understanding of polyploid plants
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