Dataset Information

Next Generation Sequencing Technologies: The Doorway to the Unexplored Genomics of Non-Model Plants.

ABSTRACT: Non-model plants i.e., the species which have one or all of the characters such as long life cycle, difficulty to grow in the laboratory or poor fecundity, have been schemed out of sequencing projects earlier, due to high running cost of Sanger sequencing. Consequently, the information about their genomics and key biological processes are inadequate. However, the advent of fast and cost effective next generation sequencing (NGS) platforms in the recent past has enabled the unearthing of certain characteristic gene structures unique to these species. It has also aided in gaining insight about mechanisms underlying processes of gene expression and secondary metabolism as well as facilitated development of genomic resources for diversity characterization, evolutionary analysis and marker assisted breeding even without prior availability of genomic sequence information. In this review we explore how different Next Gen Sequencing platforms, as well as recent advances in NGS based high throughput genotyping technologies are rewarding efforts on de-novo whole genome/transcriptome sequencing, development of genome wide sequence based markers resources for improvement of non-model crops that are less costly than phenotyping.

SUBMITTER: Unamba CI

PROVIDER: S-EPMC4679907 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Next Generation Sequencing Technologies: The Doorway to the Unexplored Genomics of Non-Model Plants.

Unamba Chibuikem I N CI Nag Akshay A Sharma Ram K RK

Frontiers in plant science 20151216

Non-model plants i.e., the species which have one or all of the characters such as long life cycle, difficulty to grow in the laboratory or poor fecundity, have been schemed out of sequencing projects earlier, due to high running cost of Sanger sequencing. Consequently, the information about their genomics and key biological processes are inadequate. However, the advent of fast and cost effective next generation sequencing (NGS) platforms in the recent past has enabled the unearthing of certain ...[more]

PMID: 26734016

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Project description:BACKGROUND: We have developed a simulation approach to help determine the optimal mixture of sequencing methods for most complete and cost effective transcriptome sequencing. We compared simulation results for traditional capillary sequencing with "Next Generation" (NG) ultra high-throughput technologies. The simulation model was parameterized using mappings of 130,000 cDNA sequence reads to the Arabidopsis genome (NCBI Accession SRA008180.19). We also generated 454-GS20 sequences and de novo assemblies for the basal eudicot California poppy (Eschscholzia californica) and the magnoliid avocado (Persea americana) using a variety of methods for cDNA synthesis. RESULTS: The Arabidopsis reads tagged more than 15,000 genes, including new splice variants and extended UTR regions. Of the total 134,791 reads (13.8 MB), 119,518 (88.7%) mapped exactly to known exons, while 1,117 (0.8%) mapped to introns, 11,524 (8.6%) spanned annotated intron/exon boundaries, and 3,066 (2.3%) extended beyond the end of annotated UTRs. Sequence-based inference of relative gene expression levels correlated significantly with microarray data. As expected, NG sequencing of normalized libraries tagged more genes than non-normalized libraries, although non-normalized libraries yielded more full-length cDNA sequences. The Arabidopsis data were used to simulate additional rounds of NG and traditional EST sequencing, and various combinations of each. Our simulations suggest a combination of FLX and Solexa sequencing for optimal transcriptome coverage at modest cost. We have also developed ESTcalc http://fgp.huck.psu.edu/NG_Sims/ngsim.pl, an online webtool, which allows users to explore the results of this study by specifying individualized costs and sequencing characteristics. CONCLUSION: NG sequencing technologies are a highly flexible set of platforms that can be scaled to suit different project goals. In terms of sequence coverage alone, the NG sequencing is a dramatic advance over capillary-based sequencing, but NG sequencing also presents significant challenges in assembly and sequence accuracy due to short read lengths, method-specific sequencing errors, and the absence of physical clones. These problems may be overcome by hybrid sequencing strategies using a mixture of sequencing methodologies, by new assemblers, and by sequencing more deeply. Sequencing and microarray outcomes from multiple experiments suggest that our simulator will be useful for guiding NG transcriptome sequencing projects in a wide range of organisms.

Dataset Information

Next Generation Sequencing Technologies: The Doorway to the Unexplored Genomics of Non-Model Plants.

Publications

Next Generation Sequencing Technologies: The Doorway to the Unexplored Genomics of Non-Model Plants.

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