A survey of the complex transcriptome from the highly polyploid sugarcane genome using full-length isoform sequencing and de novo assembly from short read sequencing

Abstract

BackgroundDespite the economic importance of sugarcane in sugar and bioenergy production, there is not yet a reference genome available. Most of the sugarcane transcriptomic studies have been based on Saccharum officinarum gene indices (SoGI), expressed sequence tags (ESTs) and de novo assembled transcript contigs from short-reads; hence knowledge of the sugarcane transcriptome is limited in relation to transcript length and number of transcript isoforms.ResultsThe sugarcane transcriptome was sequenced using PacBio isoform sequencing (Iso-Seq) of a pooled RNA sample derived from leaf, internode and root tissues, of different developmental stages, from 22 varieties, to explore the potential for capturing full-length transcript isoforms. A total of 107,598 unique transcript isoforms were obtained, representing about 71% of the total number of predicted sugarcane genes. The majority of this dataset (92%) matched the plant protein database, while just over 2% was novel transcripts, and over 2% was putative long non-coding RNAs. About 56% and 23% of total sequences were annotated against the gene ontology and KEGG pathway databases, respectively. Comparison with de novo contigs from Illumina RNA-Sequencing (RNA-Seq) of the internode samples from the same experiment and public databases showed that the Iso-Seq method recovered more full-length transcript isoforms, had a higher N50 and average length of largest 1,000 proteins; whereas a greater representation of the gene content and RNA diversity was captured in RNA-Seq. Only 62% of PacBio transcript isoforms matched 67% of de novo contigs, while the non-matched proportions were attributed to the inclusion of leaf/root tissues and the normalization in PacBio, and the representation of more gene content and RNA classes in the de novo assembly, respectively. About 69% of PacBio transcript isoforms and 41% of de novo contigs aligned with the sorghum genome, indicating the high conservation of orthologs in the genic regions of the two genomes.ConclusionsThe transcriptome dataset should contribute to improved sugarcane gene models and sugarcane protein predictions; and will serve as a reference database for analysis of transcript expression in sugarcane.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-017-3757-8) contains supplementary material, which is available to authorized users.

Allele-defined genome of the autopolyploid sugarcane Saccharum spontaneum L.

“…In addition, the RNA-seq reads were mapped to the AP85-441 genome using HiSAT2 75 version 2.10 and reassembled using StringTie 76 version 1.3.4, which is a reference-based RNA assembler. Meanwhile, published full-length transcripts based on IsoSeq in sugarcane were also recruited for annotation 77 . The four haplotypes (A, B, C and D) were split into four sub-genomes, each containing eight pseudo-molecules.…”

Transcriptome analysis based on a combination of sequencing platforms provides insights into leaf pigmentation in Acer rubrum

“…Currently, whole-transcriptome sequencing for nonmodel organisms, without reference sequences, has become an efficient tool for the extraction of useful genes and exploration of gene expression patterns. Third-generation sequencing technologies have been employed to generate highly contiguous reconstructions of many dozens of plants, such as Agaves [51], Salvia miltiorrhiza [52], rice [53], Beta vulgaris [54], Triticum aestivum [55], strawberry [56], sorghum [57], maize [58], Amborella trichopoda [59], Phyllostachys edulis [60], sugarcane [61], and Arabidopsis [62], which have provided new insights into their evolution and sequence diversity. PAC-BIO RS (Pacific Biosciences of California, Inc., http://www.…”

Machine learning approaches reveal genomic regions associated with sugarcane brown rust resistance

“…S. officinarum has a more efficient process of sugar production but is susceptible to several biotic and abiotic stresses, in contrast to S. spontaneum, which has a low sucrose content but is resistant to different types of stress 1,3,5 . Sugarcane cultivars have unique chromosome sets (with numbers ranging from 80 to 130) 6 with highly complex genomic organization 1 , a polyploid genome (with overall ploidy estimated to be between 6 and 14) 7 , a frequent occurrence of aneuploidy at the locus level depending on the number of homologous chromosomes in hybrid cultivars 8 , an estimated whole-genome size of 10 Gb 9 , and a high content of repetitive regions (50% of genome size) 10 . This complexity has challenged the efforts of the scientific community to unravel the genetic architecture of sugarcane in terms of the molecular mechanisms underlying different phenotypes, particularly efforts to detect regions of phenotype-genotype associations.…”