The genome of the Wollemi pine, a critically endangered "living fossil" unchanged since the Cretaceous, reveals extensive ancient transposon activity.

We present the genome of the living fossil, Wollemia nobilis, a southern hemisphere conifer morphologically unchanged since the Cretaceous. Presumed extinct until rediscovery in 1994, the Wollemi pine is critically endangered with less than 60 wild adults threatened by intensifying bushfires in the Blue Mountains of Australia. The 12 Gb genome is among the most contiguous large plant genomes assembled, with extremely low heterozygosity and unusual abundance of DNA transposons. Reduced representation and genome re-sequencing of individuals confirms a relictual population since the last major glacial/drying period in Australia, 120 ky BP. Small RNA and methylome sequencing reveal conservation of ancient silencing mechanisms despite the presence of thousands of active and abundant transposons, including some transferred horizontally to conifers from arthropods in the Jurassic. A retrotransposon burst 8-6 my BP coincided with population decline, possibly as an adaptation enhancing epigenetic diversity. Wollemia, like other conifers, is susceptible to Phytophthora, and a suite of defense genes, similar to those in loblolly pine, are targeted for silencing by sRNAs in leaves. The genome provides insight into the earliest seed plants, while enabling conservation efforts.

Chromosome-Level Alpaca Reference Genome VicPac3.1 Improves Genomic Insight Into the Biology of New World Camelids.

The development of high-quality chromosomally assigned reference genomes constitutes a key feature for understanding genome architecture of a species and is critical for the discovery of the genetic blueprints of traits of biological significance. South American camelids serve people in extreme environments and are important fiber and companion animals worldwide. Despite this, the alpaca reference genome lags far behind those available for other domestic species. Here we produced a chromosome-level improved reference assembly for the alpaca genome using the DNA of the same female Huacaya alpaca as in previous assemblies. We generated 190X Illumina short-read, 8X Pacific Biosciences long-read and 60X Dovetail Chicago® chromatin interaction scaffolding data for the assembly, used testis and skin RNAseq data for annotation, and cytogenetic map data for chromosomal assignments. The new assembly VicPac3.1 contains 90% of the alpaca genome in just 103 scaffolds and 76% of all scaffolds are mapped to the 36 pairs of the alpaca autosomes and the X chromosome. Preliminary annotation of the assembly predicted 22,462 coding genes and 29,337 isoforms. Comparative analysis of selected regions of the alpaca genome, such as the major histocompatibility complex (MHC), the region involved in the Minute Chromosome Syndrome (MCS) and candidate genes for high-altitude adaptations, reveal unique features of the alpaca genome. The alpaca reference genome VicPac3.1 presents a significant improvement in completeness, contiguity and accuracy over VicPac2 and is an important tool for the advancement of genomics research in all New World camelids.

Whole Genome Sequencing of the Asian Arowana (Scleropages formosus) Provides Insights into the Evolution of Ray-Finned Fishes.

The Asian arowana (Scleropages formosus) is of commercial importance, conservation concern, and is a representative of one of the oldest lineages of ray-finned fish, the Osteoglossomorpha. To add to genomic knowledge of this species and the evolution of teleosts, the genome of a Malaysian specimen of arowana was sequenced. A draft genome is presented consisting of 42,110 scaffolds with a total size of 708 Mb (2.85% gaps) representing 93.95% of core eukaryotic genes. Using a k-mer-based method, a genome size of 900 Mb was also estimated. We present an update on the phylogenomics of fishes based on a total of 27 species (23 fish species and 4 tetrapods) using 177 orthologous proteins (71,360 amino acid sites), which supports established relationships except that arowana is placed as the sister lineage to all teleost clades (Bayesian posterior probability 1.00, bootstrap replicate 93%), that evolved after the teleost genome duplication event rather than the eels (Elopomorpha). Evolutionary rates are highly heterogeneous across the tree with fishes represented by both slowly and rapidly evolving lineages. A total of 94 putative pigment genes were identified, providing the impetus for development of molecular markers associated with the spectacular colored phenotypes found within this species.

Intron splicing suppresses RNA silencing in Arabidopsis.

Silencing of introduced transgenes constitutes a major bottleneck in the production of transgenic crops. Commonly, these transgenes contain no introns, a feature shared with transposons, which are also prime targets for gene silencing. Given that introns are very common in endogenous genes but are often lacking in transgenes and transposons, we hypothesised that introns may suppress gene silencing. To investigate this, we conducted a genome-wide analysis of small RNA densities in exons from intronless versus intron-containing genes in Arabidopsis thaliana. We found that small RNA libraries are strongly enriched for exon sequences derived from intronless genes. Small RNA densities in exons of intronless genes were comparable to exons of transposable elements. To test these findings in vivo we used a transgenic reporter system to determine whether introns are able to suppress gene silencing in Arabidopsis. Introducing an intron into a transgene reduced silencing by more than fourfold. Compared with intronless transcripts, the spliced transcripts were less effective substrates for RNA-dependent RNA polymerase 6-mediated gene silencing. This intron suppression of transgene silencing requires efficient intron splicing and is dependent on ABH1, the Arabidopsis orthologue of human cap-binding protein 80.

A transcriptional sketch of a primary human breast cancer by 454 deep sequencing.

BACKGROUND: The cancer transcriptome is difficult to explore due to the heterogeneity of quantitative and qualitative changes in gene expression linked to the disease status. An increasing number of "unconventional" transcripts, such as novel isoforms, non-coding RNAs, somatic gene fusions and deletions have been associated with the tumoral state. Massively parallel sequencing techniques provide a framework for exploring the transcriptional complexity inherent to cancer with a limited laboratory and financial effort. We developed a deep sequencing and bioinformatics analysis protocol to investigate the molecular composition of a breast cancer poly(A)+ transcriptome. This method utilizes a cDNA library normalization step to diminish the representation of highly expressed transcripts and biology-oriented bioinformatic analyses to facilitate detection of rare and novel transcripts. RESULTS: We analyzed over 132,000 Roche 454 high-confidence deep sequencing reads from a primary human lobular breast cancer tissue specimen, and detected a range of unusual transcriptional events that were subsequently validated by RT-PCR in additional eight primary human breast cancer samples. We identified and validated one deletion, two novel ncRNAs (one intergenic and one intragenic), ten previously unknown or rare transcript isoforms and a novel gene fusion specific to a single primary tissue sample. We also explored the non-protein-coding portion of the breast cancer transcriptome, identifying thousands of novel non-coding transcripts and more than three hundred reads corresponding to the non-coding RNA MALAT1, which is highly expressed in many human carcinomas. CONCLUSION: Our results demonstrate that combining 454 deep sequencing with a normalization step and careful bioinformatic analysis facilitates the discovery and quantification of rare transcripts or ncRNAs, and can be used as a qualitative tool to characterize transcriptome complexity, revealing many hitherto unknown transcripts, splice isoforms, gene fusion events and ncRNAs, even at a relatively low sequence sampling.