Source: WEST VIRGINIA STATE UNIVERSITY submitted to NRP
PHENOMICS FOR CROP IMPROVEMENT: TOOLS TO CREATE NEXT GENERATION PLANT BREEDERS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
OTHER GRANTS
Reporting Frequency
Annual
Accession No.
1007397
Grant No.
2015-38821-24373
Cumulative Award Amt.
$299,940.00
Proposal No.
2015-06226
Multistate No.
(N/A)
Project Start Date
Sep 1, 2015
Project End Date
Aug 31, 2019
Grant Year
2015
Program Code
[EP]- Teaching Project
Recipient Organization
WEST VIRGINIA STATE UNIVERSITY
PO BOX 1000
INSTITUTE,WV 25112
Performing Department
Biology
Non Technical Summary
Plant phenotyping refers to high-throughput characterization of plant traits in a quantitative description for use in GWAS combined with high-throughput single nucleotide polymorphism (SNP) resources. Phenotyping refers to the holistic characterization of plant performance by multispectral image analyses via thermographic analyses of shoot and root growth with non-destructive, image-analysis-based phenotyping under controlled conditions as well as in field plots and canopies.Once our students have been exposed to modern phenotyping and GWAS techniques, they will be able to compete for jobs or pursue advanced or additional education in one of the fastest-growing areas of research and industry. We will create a plethora of opportunities for students to pursue research in phenomics and modern methods in plant breeding and will expose them to the concept of plant breeding for food security. In doing, so we will bring our students up to speed so that they can be part of the nation's strength in crop improvement.
Animal Health Component
30%
Research Effort Categories
Basic
50%
Applied
30%
Developmental
20%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
20117991081100%
Knowledge Area
201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1799 - Fiber crops, general/other;

Field Of Science
1081 - Breeding;
Goals / Objectives
Our developing understanding of the genomes and how genetic information is manifested in the phenotypes has led to dramatic advances in all areas of biology, and these advances have been the basis of new technologies in medicine, agriculture, and industry that are not yet known to many of our students. Our goal is to make it much simpler for our high school teachers to provide their students with the laboratory experiences that mirror the open-ended explorations of scientists, instead of the traditional "cookbook" labs where the students follow instructions to a predetermined result. We aim to develop lab classes that involve plant biodiversity, impact of natural products in human health in addition to the core biotechnology modules for the biology courses taught at the advanced high-school level. We plan to aim to expose students in elementary, middle and high schools with many hours of high-quality science inquiry, supported by carefully prepared science specialists. In this way, "science education" would be redefined, with a laser-sharp focus on gaining the scientific habits of mind that will be needed by everyone to successfully negotiate his or her way through our increasingly complex and confusing science higher education. To this end, we will also organize science contests that involve successful student independent experimentation. These contests will be followed by a social gathering of distinguished visitor, WVSU researchers and the students that would generate new research and collaborative opportunities at WVSU and the students will develop interests in the various fields of biology further to explore study opportunities at WVSU.The overall "program is conceived to bring together and strengthen existing expertise in phenome analyses and infuse modern phenomic technologies into a workshop that will establish a US food safety focus at WVSU. The specific objectives are described below.Objective 1: Organize special summer workshops for faculty, graduate students and undergraduate levels for extensive training in phenomics and GWAS pipelines.Objective 2: Monthly seminars by distinguished scientists from various fields in phenomics and post-seminar get-togethers
Project Methods
Tomato Analyzeris a free software (http://www.oardc.osu.edu/ vanderknaap/tomato_analyzer.htm) designed to recognize objects of a certain size and image resolution measured in dots (pixels) per inch (dpi). The software automatically determines the boundaries of a fruit in a scanned image via contour tracing, which results in a list of adjacent points describing the border of an object in an image. All fruit shape measurements are based on boundaries. The color test module "Color Test" is designed to quantify the color parameters inside the recognized boundaries. The color measurements are based on the RGB color space: R (red), G (green), and B (Blue). The average RGB values for each pixel are assigned by the Color Test module and then translated to the CIELAB color space, which uses L*, a*, b* to describe color to approximate human visual perception. The Color Test module calculates hue and chroma color descriptors based on a* and b*.A step-by-step protocol is described for digitalization of fruits and subsequent semi-automatic analysis of morphology and color attributes with the TA software package. The protocol is divided into 8 steps: 1) image collection, 2) image analysis and calibration for the Color Test, 3) manual adjustment of attributes, 4) user-defined settings, 5) saving and exporting data, and 6) examples of data analysis.Length of trichomes is measured by using ImageJ (Schneider et al., 2012). Step 1. The option "line" is chosen to create straight, segmented or freehand lines along the trichomes. Information about the line is displayed on the status bar. Double-click to alter the line width, select options "Analyse"à"Measure" to record a line length. Step 2. The ruler option is set over the trichomes. Go to Analyze → Set Scale. In the Set Scale option, the length of the line is shown in pixels. Calibrate with the known length and select units of measurement in the appropriate boxes (in our case 5 mm) and click OK. Measurements are shown using these settings. Step 3. The data for the reference scale and trichomes are measured in millimeters. To measure many trichomes, we choose AnalyzeàToolsàROI manager. Finally, data on trichome lengths are exported into Microsoft Excel.After placing the tray with roots, click the scanner icon to digitize the roots and wait a few seconds for the scan to complete and the image to appear on the screen. WinRHIZO controls the scanner directly. It is TWAIN-compatible, so it can receive images from many scanners or cameras. It can also analyze photographic images stored in TIFF or JPEG files.Analyze the roots: When the analysis is complete, roots identified by WinRHIZO are shown by colored lines in the image. Root length and diameter are measured with Regent's unique method and by Tennant's statistical method. With the former, measurements are made continuously at each point along the root. Root overlaps at forks and tips are taken into account to provide accurate measurements of length and area.With this WinRHIZO image analysis, we can quantify areas of specific colors or groups of colors and measure root morphology as a function of color. We can set the software to color various roots to be analyzed and those of the surrounding background. WinRHIZO classifies the colors present in the image into different classes before making the morphological measurements. There are many possible applications based on color analysis. This software can be used to measure leaf area (for disease and insect damage), classify seeds, measure seedling growth, and characterize mycorrhizae.

Progress 09/01/15 to 08/31/19

Outputs
Target Audience:The results of this phenomic research focus towards researchers, students and seed companies participating in genomics driven plant breeding and identification of new genes for various traits in vegetable crops like pepper and watermelon. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?We accomplished the work with our objectives and could publish our research to disseminate to lot of audience such as private companies, students, researchers working on heavy metal remediation and reclamation.We presented 8 posters in national conferences. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Kang, Sul et al. 2010Holm 1979Benjamini and Hochberg 1995). Annotation and gene ontology terms for the SNP containing sequences were identified with the WGS. Functional network analysis of candidate genes was done using the ClueGO on the cytoscape platform (Shannon, Markiel et al. 2003). The SNP data were converted to Arabidopsis homologs locus I.D. and the STRING database was used to arrange the gene network (Szklarczyk, Franceschini et al. 2010). This analyzed the relationship among the genes to perform different functions in the root under low pH. We validated our Capsicum candidates with candidates from Arabidopsis in low pH and Aluminum toxicity and 175 genes were observed to be common with all three species of Capsicum used in our study. Of these genes, the candidates with common traits were selected and the allele plot were made to observe how each allele contributed to acidity tolerance in the population we used. Their Arabidopsis homology were also identified to provide more information on the function of a particular gene and SNP.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Akinmoju T., Bodunrin A., Sharma D., Shinde S., Dianellos V., Abburi V., Ortiz C., Nimmakayala P., Saminathan T., Reddy U., Identification of Trichome Development Related Genes Using Genome-wide Association Studies in Watermelon Diverse Collections. The Donald Danforth Plant Science Center 19th Annual Fall Symposium in Saint Louis, Missouri September 26- 28, 2018
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Bhandari M., Kshetry A., Pe�a-Garc�a Y., Shinde S., Saminathan T., Nimmakayala P., Reddy U. Uncovering the Genetic Components for Acidity Tolerance in Arabidopsis by Genome Wide Association Study. 2019 Annual Meeting Midwestern Section American Society of Plant Biologists. Morgantown WV
  • Type: Journal Articles Status: Under Review Year Published: 2019 Citation: Nimmakayala P, Abburi VL, Davenport B, Nadimi M, Davidson J, Tonapi K, Yadav L, Stommel J, Reddy UK (2019) Genome-wide Diversity and Association Mapping for fruit colors in various Capsicum spp. Frontiers in Plant Science (In review)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Kshetry A., Bhandari M., Pe�a-Garc�a Y., Shinde S., Saminathan T., Nimmakayala P., Reddy U. Natural Variation in Arabidopsis thaliana to Discover Novel Genes in Response to Aluminum Toxicity. 2019 Annual Meeting Midwestern Section American Society of Plant Biologists. Morgantown WV.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Pe�a-Garc�a Y., Delgado-Ch�vez A., Lopez-Ortiz C., Balagurusamy N., Shinde S., Saminathan T., Nimmakayala P., Reddy U. Exploration into natural variation for detecting novel QTLs for Arsenic effects in Arabidopsis thaliana. ARD Research Symposium 2019. Jacksonville, FL.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Akinsipe T., Abburi L.., Nimmakayala P., Reddy U. GWAS for seedling root growth variation in acidic conditions across the cultivated pepper species complexes. ARD Research Symposium 2019. Jacksonville, FL.


Progress 09/01/17 to 08/31/18

Outputs
Target Audience: Graduate and undergraduate students of WVSU Local High school students and teachers Other 1890 faculty and graduate students Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This grant helped us to incorporate the tools and techniques developed from this grant infour graduate thesis research projects and 1 undergraduate research project. We used this teaching grant for developing capacity to perform phenomic research in watermelon, pepper and tomato. How have the results been disseminated to communities of interest?We presented 4 posters in national conferences using the phenomic resources developed in this project period. What do you plan to do during the next reporting period to accomplish the goals?We have performed several RNAseq and Genotyping by Sequencing using Illumina NextSeq® 500 next-generation sequencing (NGS) system. Graduate students and undergraduates were given intensive training to handle big datasets. During the next reporting period, we will conduct a hands on big data workshop for undergraduates. We are currently developing tools for highthroughput phenotyping anatomy of roots.

Impacts
What was accomplished under these goals? Using this teaching capacity building grant, we created several abiotic stress experiments and root phenotyping modules for teaching abiotic stress in class rooms. Students participated in phenomics experiments of root architectural changes across the stress treatments, wax related traits in various fruits, field level plant phenotyping and citrulline quantification in watermelon and used these datasets for association mapping. Students at WVSU were given metabolomic workshop in watermelon fruit flavor. We used various abiotic stress screens (arsenic and aluminum) using Arabidopsis as a model at WVSU that is being taught at several levels including Crop Evolution and Genome Level Diversity, Current Concepts of Biotechnology and Biotechniques.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Tandukar N, Saminathan T; Alaparthi S, Nimmakayala P, Hankins G and Reddy U.2018. Genome Wide Association studies on nutraceutical effects of various chili peppers on Drosophila melanogaster. 59th Annual Drosophila Research Conference, April 11-15, 2018, Philadelphia, PA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Bhandari M., Kshetry A., Shinde S., Saminathan T., Nimmakayala P., Chatfield M. and Reddy U. Elucidation of Genetic Components for Acidity and Aluminum tolerance using Arabidopsis GWAS panel. The Donald Danforth Plant Science Center 19th Annual Fall Symposium in Saint Louis, Missouri September 26- 28, 2018.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Pe�a-Garc�a Y., Delgado-Ch�vez A., Ortiz C., Balagurusamy N., Shinde S., Saminathan T., Nimmakayala P., Reddy U. Exploration into natural variation for detecting novel QTLs for Arsenic effects in Arabidopsis. The Donald Danforth Plant Science Center 19th Annual Fall Symposium in Saint Louis, Missouri September 26- 28, 2018.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Shinde S., Alvarado A., Saminathan T., Gajanayake B., Lopez C., Reddy K., Nimmakayala P., Reddy U. Analysis of miRNAome of Sweet Potato (Ipomoea batatas (L.) Lam) in response to elevated carbon dioxide and droughtThe Donald Danforth Plant Science Center 19th Annual Fall Symposium in Saint Louis, Missouri September 26- 28, 2018.


Progress 09/01/16 to 08/31/17

Outputs
Target Audience:Graduate and undergraduate students of WVSU Local High school students and teachers Other 1890 faculty and graduate students Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In addition, this grant helped us to incorporate these techniques in two graduate thesis research project and 4 undergraduate research projects. We used this teaching grant for developing capacity to perform phenomic research in watermelon, pomegranate, pepper and cotton. How have the results been disseminated to communities of interest?We presented 2 posters in national conferences using the phenomic resources developed in this project period. What do you plan to do during the next reporting period to accomplish the goals?We have recently received funding for STEM instrumentation from Department of Defense, for getting an Illumina NextSeq® 500 next-generation sequencing (NGS) system and a BioNano Genomics Irys® optical scanning platform. The instrumentation/equipment will benefit both the genomic and phenomic research and curricular environment at WVSU. It represents a considerable technological update to the Vegetable Genomics Laboratory, complementing and expanding on existing technologies used in ongoing "-omics" plant research. It will also facilitate the incorporation of genomics into WVSU's biotechnology graduate program, a significant step towards establishing a competent research and education infrastructure in phenomics and genomics at the university. We will organize another workshop for graduate students and faculty from other 1890 universities including our MSbiotech students on Nextseq technologies.

Impacts
What was accomplished under these goals? Using this teaching capacity building grant, we created comparative datasets for fruit development phenotype using automated software to digitize fruit structure and fruit color for various species of Capsicum. Students participated in phenomics experiments of fruit structure, root architectural changes across the stress treatments, fruit colors using high throughput techniques, field level plant breeding and metabolite quantification for using the datasets for association mapping. Students participating in phenomic endeavors of West Virginia State University also sequenced microbiome differences between white, red and yellow fleshed watermelons. We built genome wide association mapping pipeline at WVSU that is being taught at several levels including Crop Evolution and Genome Level Diversity, Current Concepts of Biotechnology and Biotechniques. In addition to genome assisted plant breeding, we developed several modules for trichome characterization in watermelon to be used in functional genomics. We conducted hands on workshop for WVSU and other 1890 land-grant universities in the areas of phenomics and applied plant breeding. Twenty five faculty, undergraduates and graduate students from Florida A&M University, Fort Valley State University, Alabama A&M University, Alcorn State University, Delaware State University and Tennessee State University attended a hand on workshop to learn how to use phenomic methodologies with high throughput sequencing technology to perform genomic selection for use in plant breeding.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Saminathan T, Reddy UK and Nimmakayala P. Mining trichome regulating genes in watermelon using genome wide association mapping and functional genomic approach. ASPB-Plant Biology, Minneapolis, Minnesota, USA. 2016 Jul. 26-30.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Bodunrin A, Saminathan, Nimmakayala P and Reddy UK. Genome-wide association studies (GWAS) mapping of trichome density and length in watermelon. Plant and Animal Genome XXIII  The International Conference on the Status of Plant and Animal Genome Research., San Diego, CA, USA. 2016 Jan. 10-14.


Progress 09/01/15 to 08/31/16

Outputs
Target Audience:Graduate and undergraduate students Local High school students and teachers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This grant helped us to incorporate these high-throughput techniques in one graduate thesis research project and 3 undergraduate research projects. We used this teaching grant for developing capacity to perform phenomic research in watermelon, pomegranate, pepper and cotton. How have the results been disseminated to communities of interest?Workshops, emails, conference presentations and publications. What do you plan to do during the next reporting period to accomplish the goals?We recently applied for a phenomic infrastructure grant to Department of Defence to obtain high throughput phenotyper. Using the proposal funds, we obtained WinRHIZO, a high throughput root imaging software. We will continue to incorporate phenomic modules for teaching and research endeavors in crop breeding.

Impacts
What was accomplished under these goals? Using this teaching capacity building grant, we developed various modules for phenomics using high throughput techniques, field level plant breeding and metabolite quantification for enhancing the crop improvement curriculum at West Virginia State University. We built genome wide association mapping pipeline at WVSU that is being taught at several levels including Crop Evolution and Genome Level Diversity, Current Concepts of Biotechnology and Biotechniques. In addition to genome assisted plant breeding, we developed several modules in function genomics to be used various courses. We conducted hands on workshops for WVSU and other 1890 land-grant universities in the areas of plant breeding and genomics.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Saminathan T, Bodunrin A, Singh NV, Devarajan R, Nimmakayala P, Jeff M, Aradhya M, Reddy UK (2016) Genome-wide identification of microRNAs in pomegranate (Punica granatum L.) by high-throughput sequencing. BMC Plant Biology 16 (1):1-16. doi:10.1186/s12870-016-0807-3
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Perla V, Nimmakayala P, Nadimi M, Alaparthi S, Hankins GR, Ebert AW, Reddy UK (2016) Vitamin C and reducing sugars in the world collection of Capsicum baccatum L. genotypes. Food Chemistry 202:189-198. doi:http://dx.doi.org/10.1016/j.foodchem.2016.01.135