leveraging-cyverse-resources-for-de-novo-comparative-transcriptomics

Leveraging CyVerse Resources for <em>De Novo</em> Comparative Transcriptomics of Underserved (Non-model) Organisms

This workflow allows novice researchers to leverage advanced computational resources such as cloud computing to carry out pairwise comparative ...

University of Arizona

Meiotic recombination in eukaryotes requires two homologues of the E. coli RecA proteins: Rad51 and Dmc1. Both proteins play important roles in the binding of single stranded DNA, homology search, strand invasion and strand exchange. Meiotic recombination has been well studied in Arabidopsis, rice, maize and the orthologues of RAD51 and DMC1 have been characterized. However genetic analysis of the RAD51 and DMC1 genes in bread wheat has been hampered due to the absence of complete sequence information and because of the existence of multiple copies of each gene in the hexaploid wheat genome.

The RAD51 and DMC1 homoeologous genes of bread wheat: cloning, molecular characterization and expression analysis.

Although applied over extremely short timescales, artificial selection has dramatically altered the form, physiology, and life history of cultivated plants. We have used RNAseq to define both gene sequence and expression divergence between cultivated tomato and five related wild species. Based on sequence differences, we detect footprints of positive selection in over 50 genes. We also document thousands of shifts in gene-expression level, many of which resulted from changes in selection pressure. These rapidly evolving genes are commonly associated with environmental response and stress tolerance. The importance of environmental inputs during evolution of gene expression is further highlighted by large-scale alteration of the light response coexpression network between wild and cultivated accessions. Human manipulation of the genome has heavily impacted the tomato transcriptome through directed admixture and by indirectly favoring nonsynonymous over synonymous substitutions. Taken together, our results shed light on the pervasive effects artificial and natural selection have had on the transcriptomes of tomato and its wild relatives.

Comparative transcriptomics reveals patterns of selection in domesticated and wild tomato.

The mapping and functional analysis of quantitative traits in Brassica rapa can be greatly improved with the availability of physically positioned, gene-based genetic markers and accurate genome annotation. In this study, deep transcriptome RNA sequencing (RNA-Seq) of Brassica rapa was undertaken with two objectives: SNP detection and improved transcriptome annotation. We performed SNP detection on two varieties that are parents of a mapping population to aid in development of a marker system for this population and subsequent development of high-resolution genetic map. An improved Brassica rapa transcriptome was constructed to detect novel transcripts and to improve the current genome annotation. This is useful for accurate mRNA abundance and detection of expression QTL (eQTLs) in mapping populations. Deep RNA-Seq of two Brassica rapa genotypes-R500 (var. trilocularis, Yellow Sarson) and IMB211 (a rapid cycling variety)-using eight different tissues (root, internode, leaf, petiole, apical meristem, floral meristem, silique, and seedling) grown across three different environments (growth chamber, greenhouse and field) and under two different treatments (simulated sun and simulated shade) generated 2.3 billion high-quality Illumina reads. A total of 330,995 SNPs were identified in transcribed regions between the two genotypes with an average frequency of one SNP in every 200 bases. The deep RNA-Seq reassembled Brassica rapa transcriptome identified 44,239 protein-coding genes. Compared with current gene models of B. rapa, we detected 3537 novel transcripts, 23,754 gene models had structural modifications, and 3655 annotated proteins changed. Gaps in the current genome assembly of B. rapa are highlighted by our identification of 780 unmapped transcripts. All the SNPs, annotations, and predicted transcripts can be viewed at http://phytonetworks.ucdavis.edu/.

Polymorphism identification and improved genome annotation of Brassica rapa through Deep RNA sequencing.

Shade from neighboring plants limits light for photosynthesis; as a consequence, plants have a variety of strategies to avoid canopy shade and compete with their neighbors for light. Collectively the response to foliar shade is called the shade avoidance syndrome (SAS). The SAS includes elongation of a variety of organs, acceleration of flowering time, and additional physiological responses, which are seen throughout the plant life cycle. However, current mechanistic knowledge is mainly limited to shade-induced elongation of seedlings. Here we use phenotypic profiling of seedling, leaf, and flowering time traits to untangle complex SAS networks. We used over-representation analysis (ORA) of shade-responsive genes, combined with previous annotation, to logically select 59 known and candidate novel mutants for phenotyping. Our analysis reveals shared and separate pathways for each shade avoidance response. In particular, auxin pathway components were required for shade avoidance responses in hypocotyl, petiole, and flowering time, whereas jasmonic acid pathway components were only required for petiole and flowering time responses. Our phenotypic profiling allowed discovery of seventeen novel shade avoidance mutants. Our results demonstrate that logical selection of mutants increased success of phenotypic profiling to dissect complex traits and discover novel components.

Shade avoidance components and pathways in adult plants revealed by phenotypic profiling.

Docker has become a very popular container-based virtualization platform for software distribution that has revolutionized the way in which scientific software and software dependencies (software stacks) can be packaged, distributed, and deployed. Docker makes the complex and time-consuming installation procedures needed for scientific software a one-time process. Because it enables platform-independent installation, versioning of software environments, and easy redeployment and reproducibility, Docker is an ideal candidate for the deployment of identical software stacks on different compute environments such as XSEDE and Amazon AWS. CyVerse's Discovery Environment also uses Docker for integrating its powerful, community-recommended software tools into CyVerse's production environment for public use. This paper will help users bring their tools into CyVerse Discovery Environment (DE) which will not only allows users to integrate their tools with relative ease compared to the earlier method of tool deployment in DE but will also help users to share their apps with collaborators and release them for public use.

Article	Year
The RAD51 and DMC1 homoeologous genes of bread wheat: cloning, molecular characterization and expression analysis. BMC research notes\| PubMed ID: 20920212	2010
Comparative transcriptomics reveals patterns of selection in domesticated and wild tomato. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 23803858	2013
Polymorphism identification and improved genome annotation of Brassica rapa through Deep RNA sequencing. G3 (Bethesda, Md.)\| PubMed ID: 25122667	2014
Shade avoidance components and pathways in adult plants revealed by phenotypic profiling. PLoS genetics\| PubMed ID: 25874869	2015
Bringing your tools to CyVerse Discovery Environment using Docker. F1000Research\| PubMed ID: 27803802	2016

Upendra Kumar Devisetty

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