RNA-Seq Analysis of Cocos nucifera: Transcriptome Sequencing and De Novo Assembly for Subsequent Functional Genomics Approaches

Abstract

Background Cocos nucifera (coconut), a member of the Arecaceae family, is an economically important woody palm grown in tropical regions. Despite its agronomic importance, previous germplasm assessment studies have relied solely on morphological and agronomical traits. Molecular biology techniques have been scarcely used in assessment of genetic resources and for improvement of important agronomic and quality traits in Cocos nucifera, mostly due to the absence of available sequence information.Methodology/Principal FindingsTo provide basic information for molecular breeding and further molecular biological analysis in Cocos nucifera, we applied RNA-seq technology and de novo assembly to gain a global overview of the Cocos nucifera transcriptome from mixed tissue samples. Using Illumina sequencing, we obtained 54.9 million short reads and conducted de novo assembly to obtain 57,304 unigenes with an average length of 752 base pairs. Sequence comparison between assembled unigenes and released cDNA sequences of Cocos nucifera and Elaeis guineensis indicated that the assembled sequences were of high quality. Approximately 99.9% of unigenes were novel compared to the released coconut EST sequences. Using BLASTX, 68.2% of unigenes were successfully annotated based on the Genbank non-redundant (Nr) protein database. The annotated unigenes were then further classified using the Gene Ontology (GO), Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases.Conclusions/SignificanceOur study provides a large quantity of novel genetic information for Cocos nucifera. This information will act as a valuable resource for further molecular genetic studies and breeding in coconut, as well as for isolation and characterization of functional genes involved in different biochemical pathways in this important tropical crop species.

De Novo Assembly and Analysis of the Testes Transcriptome from the Menhaden, Bervoortia tyrannus

“…Despite the importance of this fish, detailed genetic information for it is currently lacking. Transcriptome analyses are a cost-effective method for the characterization of species that lack a fully sequenced genome [7,8]. The short reads from Illumina paired-end sequencing can provide a deep sequencing coverage for the accurate base calling in de-novo transcriptome assembly and has been utilized to characterize many non-model species [37][38][39][40].…”

“…This can be investigated with either in silico or in vitro approaches. One in silico approach is to match the contigs to known cDNA or EST sequences to confirm the accuracy of the de novo assembly [8,46]. However, due to the scarcity of cDNA or EST sequences for menhaden, another alignment technique was performed for the validation.…”

“…The first is the reduction fishery that turns the menhaden into fish oil omega-3 supplements for example, and into fishmeal for livestock and aquaculture consumption [4,5]. The second is the bait fishery that uses the menhaden as bait for bluefish, crab, and lobster [6][7][8][9][10][11][12][13][14][15]. They have been rarely studied at the genetic level most likely due to the lack of genomic and transcriptome data.…”

Harnessing Next Generation Sequencing in Climate Change: RNA-Seq Analysis of Heat Stress-Responsive Genes in Wheat (Triticum aestivum L.)

“…RNA-Seq is virtually changing the area of sequencing and transcriptome profiling (Wang et al, 2009). Recently, de novo assembly has been used in some of the agriculturally important crops like chilli pepper , coconut (Fan et al, 2013), rice (Zhang et al, 2013), and sugarcane (Cardoso-Silva et al, 2014) for identification of novel genes. We used Illumina HiSeqÔ 2000 and Roche GS-FLX 454 for high-throughput deep sequencing of a heat-sensitive wheat cv.…”

Comparative transcriptomic analysis reveals a series of single nucleotide polymorphism between red- and white-fleshed loquats (Eriobotrya japonica)

“…De novo assembly of the loquat transcriptome from the fruit tissues was performed using Trinity, a short-read assembly program, as described in Haas et al (2013). The annotation for protein-coding unigene was done as follows: unigenes were compared using BLAST against the Nr database (NCBI non-redundant sequence database) using an E-value cut-off of 10 -5 and then against other protein databases, such as Swiss-Prot, Clusters of Orthologous Groups (COG) and the Kyoto Encyclopaedia of Genes and Genomes (KEGG), following the method used by Fan et al (2013).…”