Organelle Genomes and Transcriptomes of Nymphaea Reveal the Interplay between Intron Splicing and RNA Editing.

Posttranscriptional modifications, including intron splicing and RNA editing, are common processes during regulation of gene expression in plant organelle genomes. However, the intermediate products of intron-splicing, and the interplay between intron-splicing and RNA-editing were not well studied. Most organelle transcriptome analyses were based on the Illumina short reads which were unable to capture the full spectrum of transcript intermediates within an organelle. To fully investigate the intermediates during intron splicing and the underlying relationships with RNA editing, we used PacBio DNA-seq and Iso-seq, together with Illumina short reads genome and transcriptome sequencing data to assemble the chloroplast and mitochondrial genomes of Nymphaea 'Joey Tomocik' and analyze their posttranscriptional features. With the direct evidence from Iso-seq, multiple intermediates partially or fully intron-spliced were observed, and we also found that both cis- and trans-splicing introns were spliced randomly. Moreover, by using rRNA-depleted and non-Oligo(dT)-enrichment strand-specific RNA-seq data and combining direct SNP-calling and transcript-mapping methods, we identified 98 and 865 RNA-editing sites in the plastome and mitogenome of N. 'Joey Tomocik', respectively. The target codon preference, the tendency of increasing protein hydrophobicity, and the bias distribution of editing sites are similar in both organelles, suggesting their common evolutionary origin and shared editing machinery. The distribution of RNA editing sites also implies that the RNA editing sites in the intron and exon regions may splice synchronously, except those exonic sites adjacent to intron which could only be edited after being intron-spliced. Our study provides solid evidence for the multiple intermediates co-existing during intron-splicing and their interplay with RNA editing in organelle genomes of a basal angiosperm.

High-quality Cymbidium mannii genome and multifaceted regulation of crassulacean acid metabolism in epiphytes.

Epiphytes with crassulacean acid metabolism (CAM) photosynthesis are widespread among vascular plants, and repeated evolution of CAM photosynthesis is a key innovation for micro-ecosystem adaptation. However, we lack a complete understanding of the molecular regulation of CAM photosynthesis in epiphytes. Here, we report a high-quality chromosome-level genome assembly of a CAM epiphyte, Cymbidium mannii (Orchidaceae). The 2.88-Gb orchid genome with a contig N50 of 22.7 Mb and 27 192 annotated genes was organized into 20 pseudochromosomes, 82.8% of which consisted of repetitive elements. Recent expansions of long terminal repeat retrotransposon families have made a major contribution to the evolution of genome size in Cymbidium orchids. We reveal a holistic scenario of molecular regulation of metabolic physiology using high-resolution transcriptomics, proteomics, and metabolomics data collected across a CAM diel cycle. Patterns of rhythmically oscillating metabolites, especially CAM-related products, reveal circadian rhythmicity in metabolite accumulation in epiphytes. Genome-wide analysis of transcript and protein level regulation revealed phase shifts during the multifaceted regulation of circadian metabolism. Notably, we observed diurnal expression of several core CAM genes (especially ßCA and PPC) that may be involved in temporal fixation of carbon sources. Our study provides a valuable resource for investigating post-transcription and translation scenarios in C. mannii, an Orchidaceae model for understanding the evolution of innovative traits in epiphytes.

Genome skimming herbarium specimens for DNA barcoding and phylogenomics.

BACKGROUND: The world's herbaria contain millions of specimens, collected and named by thousands of researchers, over hundreds of years. However, this treasure has remained largely inaccessible to genetic studies, because of both generally limited success of DNA extraction and the challenges associated with PCR-amplifying highly degraded DNA. In today's next-generation sequencing world, opportunities and prospects for historical DNA have changed dramatically, as most NGS methods are actually designed for taking short fragmented DNA molecules as templates. RESULTS: As a practical test of routine recovery of rDNA and plastid genome sequences from herbarium specimens, we sequenced 25 herbarium specimens up to 80 years old from 16 different Angiosperm families. Paired-end reads were generated, yielding successful plastid genome assemblies for 23 species and nuclear rDNAs for 24 species, respectively. These data showed that genome skimming can be used to generate genomic information from herbarium specimens as old as 80 years and using as little as 500 pg of degraded starting DNA. CONCLUSIONS: The routine plastome sequencing from herbarium specimens is feasible and cost-effective (compare with Sanger sequencing or plastome-enrichment approaches), and can be performed with limited sample destruction.