Genomics
View abstracts in the conference PDF booklet.
Exploring Genetic Diversity in the U.S. National Melon Collection
Kaori Ando¹, Xin Wang², Umesh Reddy³, Zhangjun Fei², James D. McCreight¹
¹U.S. Department of Agriculture, Agricultural Research Service, Crop Improvement and Protection Research Unit, Salinas, CA, USA. ²Boyce Thompson Institute, Ithaca, NY, USA. ³Gus R. Douglass Institute and Department of Biology, West Virginia State University, Institute, WV, USA
It is essential to maintain, as well as to incorporate genetic diversity into breeding programs for the improvement of quality, nutrients, yield, and biotic and abiotic stress tolerances. The U.S. National Plant Germplasm System (NPGS) maintains more than 2K Cucumis melo L. accessions from >70 countries. The NPGS melon collection has a paucity of systematically collected phenotypic data, and few of them have been classified to the horticultural Group level. These accessions and additional heirloom cultivars not in the NPGS collection were genotyped by the Genotyping by Sequence (GBS) method to understand the genetic diversity, population structure, and phylogenetic relationships within the U.S. melon collection. Over 27K single nucleotide polymorphism (SNP) markers with missing rate less than 0.5 and minimum allele frequency greater than 0.01 were obtained. Eleven sub-populations were identified via population structure analysis. We then developed a core panel subset (n=384) based on genome-wide SNPs, origin and known phenotypic properties, which captured 98.96% of the allelic diversity of the base population. The core panel was evaluated in a non-replicated field test in Imperial Valley, CA for 10 phenotypic traits (leaf shape, sex expression, and fruit traits including exocarp, flesh and cavity color, shape, weight, and soluble solids). Each member of the core panel was classified for horticultural Group based on phenotypic data, and we examined the relationship between genotypic data and horticultural Group classification. The core panel data were subjected to genome-wide association study in order to identify marker- trait associations for the key phenotypic traits. Overall, characterization of the genetic diversity and structure of the entire U.S. melon collection based on the high-density SNP markers derived from GBS and establishment of a melon core panel provide a valuable resource for melon genetic improvement. Partial Funding provided by the Specialty Crop Block Grant Program at the U.S. Department of Agriculture (USDA) through Grant 14-SCBGP-CA-0006 (The CucCAP Project)
Transcriptome Methods Identified Mobile mRNAs from Pumpkin Rootstock in Watermelon Scions that Respond to Chilling Stress
Fei Cheng, Junyang Lu, Nannan Wei, Zhilong Bie Huazhong
Agricultural University, Wuhan, China
Grafting of scion onto resistant pumpkin rootstock is an effective method to enhance chilling tolerance of watermelon seedlings. Previous studies showed that grafting watermelon scion onto rootstock led to differential expression of watermelon genes. However, the regulatory mechanisms remain to be investigated. In this study, the RNA-sequence technique was used to analyze the mechanism of mobile mRNAs from pumpkin rootstock to regulate chilling tolerance in watermelon. The leaves of self-grafted and rootstock-grafted watermelon were respectively collected at 12 h and 72 h from control (28/18 °C) and chilling group (10/10 °C), and were used for building library and sequencing. The results showed the detection of 91, 73, 97, and 100 mobile mRNAs in the treatment groups of control-12 h, control-72 h, chilling-12 h, and chilling- 72 h from the pumpkin rootstock, respectively. Gene Ontology (GO) analysis of these mobile mRNAs indicated that certain biological processes, such as metabolic process (64.6 %), cellular (63.5 %), single-organism process (49.0 %), were overrepresented. Within the cellular component category, cell (66.1 %), cell part (66.1 %), and organelle (60.6 %) were overrepresented. Moreover, binding (65.5 %) and catalytic activity (49.0 %) were overrepresented in the molecular function category. After 12 hours of chilling stress, total 46 specific mRNAs were detected to be involved in long-distance transportation, and the number was increased to 83 at 72 hours of chilling stress. These results provide further insights into the molecular mechanisms of pumpkin rootstock grafting to increase chilling tolerance of watermelon seedlings, and can be helpful to identify some suitable candidate genes that conveying valuable information for improving chilling tolerance in watermelon through genetic engineering.
Cucurbit Genomics Database
Zhangjun Fei
Boyce Thompson Institute, Ithaca, NY, USA
The Cucurbitaceae family (cucurbit) includes several economically important crops, such as melon, cucumber, watermelon, pumpkin, squash, and gourd. During the past several years, genomic and genetic data have been rapidly accumulated for cucurbits. We have been developing the Cucurbit Genomics Database (http://cucurbitgenomics.org/) to store, mine, analyze, and disseminate the large-scale cucurbit genomic and genetic datasets in an efficient way and to provide a center portal for the cucurbit research and breeding community. The database currently contains all available genome and EST sequences, genetic maps, and transcriptome profiling for cucurbit species, as well as sequence annotations and biochemical pathways. A set of analysis and visualization tools and user-friendly query interfaces have been implemented in the database. Future development of the database will be discussed.
Mapping of QTLs Controlling Seed Size by Whole-Genome Sequencing and Bulk Segregation Analysis in Watermelon
Meiling Gao¹,², Yu Guo¹, Xiujie Liu³, Jixiu Liu³, Yanling Zhang¹, Xiaoxue Liang¹
¹College of Life Sciences, Agriculture and Forestry,Qiqihar University, Qiqihar, 161006, China. ²Heilongjiang Provincial Key Laboratory of Resistance Gene Engineering and Protection of Biodiversity in Cold Areas, Qiqihar, China. ³Qiqihar Horticultural Research Institute, Qiqihar, China
Seed size controlled by QTLs dispersed in watermelon genome is a quantitative trait that influences both the quality and yield of watermelon. QTL-seq a rapid high-resolution genome- wide strategy , combining bulked segregation analysis (BSA) with whole genome sequencing , is the first steps for further research, such as cloning and function analysis . In this study, an F2 population derived from a cross between a inbred line K1 which the seed length was 5.8 mm and a inbred line L1 which the seed length was 9.8 mm, was used for excavation of controlling seed size traits genes by BSA-seq. The “L-Bulk” and “S-Bulk” DNA bulks were constructed using 20 plants selected from the F2 population. Next-generation sequencing (NGS) was applied for the resequencing of the parental bulks and two bulks . The result of the BSA showed a major region on chromosome 6 was identified about 2.82 Mb. Biparental QTL was conducted in the candidate region. A F2 genetic linkage map which included 21 CAPS markers and 1 dCAPS markers was constructed. The map contained 1 linkage groups which corresponded with the chromosome and spanned 60.03 cM with a mean marker interval of 2.73 cM, a major-effect QTL named SS6.1 was detected between the markers WSS-18 and CAPS-29 which was tightly linked to the seed length and 100 seed weight. The QTL analysis indicated the presence of one quantitative trait loci, SS6.1 was found in about 460 kb region WSS-18 and CAPS-29, which mapped on chromosome 6 of watermelon genome. There were 39 candidate genes were predicted in this region. Based on the watermelon genome and function analysis, five genes including Cla009263, Cla009266, Cla009289, Cla009292, Cla009303 were predicted as candidate genes related to the seed size. The results of candidate gene analysis provided some potential target for further cloning and functional identification of the seed size for breeding.
The CucCAP Project: Genomic Tools and Resources to Facilitate Breeding for Disease Resistance in Cucurbits
Rebecca Grumet¹, Zhangjun Fei², Yiqun Weng³, Xin Wang², Kan Bao², Yi Zheng², Todd Wehner4, Umesh Reddy5, Amnon Levi6, James D. McCreight7, Michael Mazourek8, Shaker Kousik6, Kai-Shu Ling6, Cecilia McGregor9, W. Patrick Wechter6, Linda Wessel-Beaver10, William M. Wintermantel7, Mary Hausbeck1, Angela Linares-Ramirez10, Lina Quesada-Ocampo4, Christine Smart8
¹Michigan State University, East Lansing, MI, USA. ²Boyce Thompson Institute, Ithaca, NY, USA. ³U.S. Department of Agriculture, Agricultural Research Service, Vegetable Crops Research Unit, Madison, Madison, WI, USA. 4North Carolina State University, Raleigh, NC, USA. 5West Virginia State University, Institution, WV, USA. 6U.S. Department of Agriculture, Agricultural Research Service, Charleston, SC, USA. 7U.S. Department of Agriculture, Agricultural Research Service, Crop Improvement and Protection Research Unit, Salinas, CA, USA. 8Cornell University, Ithaca, NY, USA. 9University of Georgia, Athens, GA, USA. 10University of Puerto Rico, Mayaguez, PR, USA
The USDA-SCRI ‘CucCAP’ project is an effort by the U.S. cucurbit community to develop genomic and bioinformatic tools for the Cucurbitaceae to facilitate introgression and stacking of disease resistance loci. Tool development includes upgrading of the Cucurbit Genomics Database (http://cucurbitgenomics.org/) which provides sequence data, search capacity, and bioinformatics tools, and genetic characterization of the National Plant Germplasm System (NPGS) plant introduction (PI) collections of watermelon (Citrullus lanatus), melon (Cucumis melo), cucumber (Cucumis sativus), and squash (Cucurbita pepo). Genotyping-by-sequencing (GBS) was performed on the full collections (1000-2000 accessions/crop) providing 0.9-1.7 billion GBS reads and 20,000-30,000 SNPs per species that are well distributed across the genomes (average density: one SNP per 10.6, 14.6, and 15.7 kb for cucumber, melon and watermelon, respectively). The SNPs were used to characterize genetic diversity, population structure, phylogenetic relationships, linkage disequilibrium, and population differentiation of the collections and perform GWAS of horticulturally important traits. This information is being used to establish publicly available, re-sequenced functional panels of 300-400 accessions per crop representing >95% of the diversity present in the collections along with key disease resistance, fruit quality, horticultural and agronomic traits. The CucCAP disease priorities, which were identified by the cucurbit industries and vary among crops, include downy mildew, Fusarium, gummy stem blight, Phytophthora capsici, powdery mildew and several viruses. The participating research groups are characterizing resistances, identifying QTL, developing markers, and introgressing resistances for the different crop-disease combinations. Partial Funding provided by the Specialty Crop Block Grant Program at the U.S. Department of Agriculture (USDA) through Grant 14-SCBGP-CA-0006 (The CucCAP Project)
Quality and Weight
Marleny Lozano1, Hugh Dalton1, Sudip Dutta1, Yan Tomason1, Carlos Ortiz1, Padma Nimmakayala1, Amnon Levi2
1West Virginia State University, Institute, WV, USA. 2U.S. Department of Agriculture, Agricultural Research Service, U.S. Vegetable Laboratory, Charleston, SC, USA
To assess the impact of grafting on gene expression in fruits, we compared transcriptome generated using RNAseq for fruit tissues collected in scions and ungrafted plants of watermelon and Lagenaria (bottle gourd). When watermelon is used as scion (WM/LAG), 57 genes were shown differentially expressed (DEGs), when compared to non-grafted watermelon of which, 11 were upregulated and 46 were downregulated. Four genes were exclusively absent in WM/LAG while they highly expressed in nongrafted watermelon. Upregulated genes were of plant cell wall modification, carbohydrate metabolism, synthesis of volatile compounds, hormone metabolism and stress response. Pectinesterase (Cla004251), 5-dehydro-2- deoxygluconokinase (Cla007008) germacrene D synthase (Cla004416), protein forked1 (Cla021595) and phloem filament protein (Cla001440) were examples for upregulated DEGs. On the other hand, downregulated genes were associated to transport, response to stimulus, transcription factors, signaling and metabolic process including peptide transporter (Cla018169), calcium-dependent lipid binding protein (Cla006159), ethylene responsive transcription factor (Cla014156), elicitor-responsive protein (Cla016038) and biosynthetic arginine decarboxylase (Cla016612). 478 genes were significantly differentially expressed in fruits of LAG/WM when compared with nongrafted bottle gourd. Of these genes, 263 were upregulated and 215 were downregulated. Hormone metabolism, transporter activity, response to stimulus and oxidative stress were found among the DEGs. ATP-dependent 6- phosphofructokinase (Lsi07G005330), auxin efflux carrier family protein (Lsi01G016410), ethylene-responsive transcription factor (Lsi06G008160), high-affinity glucose transporter (Lsi09G013320), potassium transporter (Lsi02G015540), calcium-binding EF hand protein (Lsi02G010860) and amine oxidase (Lsi09G013240) were some of the notable DEGs in the fruits of LAG/WM. In this study, we also field evaluated reciprocal grafts to understand the effects of grafting on fruits.
The Whole-Genome Resequencing Reveals the Evidence of Selective Sweeps during Melon Domestication
Shi Liu, Peng Gap, Qianglong Zhu, Xuezheng Wang, Hongyu Liu, Zicheng Zhu, Hongyan Ma, Feishi Luan
College of Horticulture and Landscape Architecture, Northeast Agricultural University, Harbin, Heilongjiang, China
As an important member of cucurbitaceae family, melon exhibits the most diversity in phenotype. However, the genomic evidence of melon domestication remains poorly understood. The goal of this study is to investigate the genomic signatures in the history of melon domestication. We resequenced 294 melon accessions to get an ultra-genomic variation map with 2,045,412 SNPs. Genomic FST analysis of wild versus domesticated populations suggests that genes which affect aroma, sweetness, flower time and stress resistance have undergone strong positive selection during domestication. Based on the results of the domesticated signal selection, we speculated that the evolution in melon from wildtype to modern cultivated species underwent two stages (the domestication stage and the improvement stage). The first one is the domestication from the wildtype to two kinds of landraces closely to the C. melo ssp. melo and C. melo ssp. agrestis with different selective sweeps (203 and 178 candidate genes in the selective regions with the 1% top FST values, respectively). For the landrace closely to the C. melo ssp. agrestis, the genes related to sugar accumulation and disease-resistance were selective, while in the C. melo ssp. melo, the stress tolerance and plant development genes performed high selective signals. The aroma related genes were selected in both landraces. The two kinds of landraces might have undergone independent evolution to the varieties of C. melo ssp. melo and C. melo ssp. agrestis, respectively in the improved stages. In the second stages, the C. melo ssp. agrestis underwent much more genome variation than the C. melo ssp. melo to perform a high degree domestication events. The C. melo ssp. melo and C. melo ssp. agrestis groups did not have any domesticated relationships in the evolutionary history. This study advances the understanding of melon domestication and also gives the genome signals to enhance the melon breeding.
Metagenomic and Metatranscriptomic Analyses of Diverse Watermelon Cultivars Reveal the Role of Fruit-Associated Microbiome in Carbohydrate Metabolism and Ripening of Mature Fruits
Carlos Ortiz1, Umesh Reddy1, Marleny Garcia1, Thangasamy Saminathan1, Padma Nimmakayala1, Nagamani Balagurusamy2
1West Virginia State University, Institute, USA. 2Universidad Autónoma de Coahuila, Torreón, Mexico
The plant microbiome is a key determinant of plant health and productivity, and changes in the plant microbiome can alter the tolerance to biotic and abiotic stress and the quality of end produce. In this study, we aimed to understand the diversity and function of microorganisms in relation to ripening and carbohydrate metabolism in ripe watermelon fruits. We used 16S metagenomics and RNAseq metatranscriptomics for analysis of red (PI 459074, ‘Congo’ and SDRose) and yellow (PI 227202, PI 435990 and ‘JBush’) flesh watermelon germplasm of geographically and metabolically diverse origin. Metagenomics data showed that proteobacteria were abundant in Sweet Dakota Rose (SDRose) and PI227202, whereas cyanobacteria were most abundant in Congo and PI4559074. In the case of metatranscriptome data, Proteobacteria was the most abundant in all cultivars. High expression of genes linked to infectious diseases and the expression of peptidoglycan hydrolases associated to pathogenicity of eukaryotic hosts was observed in SDRose, which could have resulted in low microbial diversity in this cultivar. Production of carotenoids and other volatile aroma compounds due to carotenoid degradation could be correlated with presence of active microbial groups Cyanobacteria and Deinococcus-Thermus. Moreover, Basidiomycota and Ascomycota also could contribute to volatile aroma compounds and their activity is observed in the metatranscriptomic results. Further, ethylene production activity leading to fruit ripening could be associated with Proteobacteria. The presence of GH28, associated with polygalacturonase activity in JBush and SDRose could be related to cell wall modifications including de- esterification and depolymerization, and consequent loss of galacturonic acid and neutral sugars. Moreover, based on the KEGG annotation of the expressed genes, nine α-galactosidase genes involved in key processes of galactosyl oligosaccharide metabolism, such as raffinose family oligosaccharide were identified and galactose metabolism pathway was reconstructed. Results of this study underline the links between the host and fruit-associated microbiome in carbohydrate metabolism of the host and fruit ripening. The cultivar difference in watermelon reflects the quantum and diversity of the microbiome, which would benefit watermelon and other plant breeders aiming at the holobiont concept to incorporate associated microbiomes in breeding plan.
A Multispecies SNP Array for High-Resolution Genotyping of Melon, Cucumber and Watermelon
Martin Ganal, Andreas Polley, Joerg Plieske, Eva Maria Graner
TraitGenetics GmbH, Stadt Seeland OT Gatersleben, 06466, Germany
With the availability of a set of sequenced cucumber, watermelon and melon genomes in public databases, a large source of SNPs is now available for these important Cucurbitaceae species. We have used these resources of molecular markers for the development of a new multispecies SNP array using the Affymetrix Axiom genotyping platform. The array has been set up with 29,961 melon, 47,537 watermelon, and 49,497 cucumber SNP markers. For each species, the markers have been selected based on (i) general allele frequency in all sequenced lines; (ii) chromosomal distribution along the physical length of the chromosomes with a higher marker number towards the end of the chromosomes reflecting the distribution of crossing overs; and (iii) expected marker functionality over a wide range of material. This new Cucurbitaceae SNP genotyping array has been used for the characterization of sets of cucumber, melon and watermelon breeding lines and varieties for defining the individual marker functionality and quality as well as the general level of polymorphism. With many functional markers, this SNP genotyping array provides a significant improvement for large-scale genotyping in these Cucurbitaceae species.
New Genetic and Genomic Resources in Melon and their Application for Fruit Quality Improvement
Valentino Ruggieri1,2, Lara Pereira1, Konstantinos Alexiou1,2, Jordi Morata1, Jason Argyris1,2, Sergi Pérez1, Riccardo Aiese Cigliano3, Walter Sanseverino3, Pere Puigdomènech1, Josep Casacuberta1, Jordi Garcia-Mas1,2, Marta Pujol1,2
1Centre for Research in Agricultural Genomics (CRAG) CSIC-IRTA-UAB-UB, Bellaterra, Barcelona, Spain. 2IRTA (Institut de Recerca i Tecnologia Agroalimentàries), Bellaterra, Barcelona, Spain. 3Sequentia Biotech SL, Bellaterra, Barcelona, Spain
During the last years, a big effort has been done in order to develop genetic and genomic tools in melon (Cucumis melo L.) that are useful to identify the genes or QTLs underlying traits of high agronomical importance. In this sense, we have improved the assembly of the melon reference genome (v3.6.1, http://melonomics.net) by using an optical mapping approach, obtaining the correct order and orientation of 21 scaffolds, and defining the gap-size in the 12 pseudomolecules. A new annotation v4.0 integrating RNA-seq data has been released, with more than 8,000 new genes identified. These resources are very useful to identify candidate genes obtained from mapping experiments. With the aim to improve melon fruit quality, we developed a RIL population between two commercial cultivars: the highly climacteric “Védrantais” from the cantalupensis group, and the non-climacteric “Piel de Sapo” from the inodorus group. This population segregates for many fruit quality and morphology traits, and we used a GBS strategy to map QTLs for sugar and carotenoid content, fruit and seed morphology, and the external appearance of the fruit. The high mapping resolution allowed the identification of five major loci, and 33 QTLs in intervals of 1 Mb containing 100 genes on average.
Genomics-Aided Development and Characterization of Cucumis hystrix Introgression Lines in Cucumber
Paradee Thammapichai1, Junsong Pan1,2, Dal-Hoe Koo1, Yonghua Han1,3, Jiming Jiang1, Yiqun Weng4
1University of Wisconsin, Madison, WI, USA. 2Department of Plant Science, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, China. 3The Key Laboratory of Biotechnology for Medicinal Plants of Jiangsu Province, Jiangsu Normal University, Xuzhou, China. 4U.S. Department of Agriculture, Agricultural Research Service, Vegetable Crops Research Unit, Madison, WI, USA
Cucumis hystrix (2n = 2x = 24, HH) is the only known species in the genus Cucumis that is cross- compatible with cucumber (C. sativus, 2n = 2x = 14, CC) and has a great potential for cucumber improvement. To facilitate introgression of C. hystrix chromatins into cucumber genetic background, we developed a draft genome for the C. hystrix genome (accession TH1) which contained 16,865 scaffolds (~78× coverage) for a total of 226.0 Mb, representing ~51% of the estimated 447 Mb C. hystrix genome. The largest scaffold was 342 kb with the N50 scaffold size of 23.3 kb. Through genotyping-by-sequencing (GBS), a linkage map for C. hystrix was developed with 1,692 SNP loci, which was then integrated with a previously developed genetic map with 410 SSR markers. The resulting consensus map consisted of 12 linkage groups spanning 1119 cM, which was used to anchor 1,069 scaffolds accounting for 49.4 Mb or ~22% of C. hystrix draft genome assembly. A karyotype for the C. hystrix genome was developed with molecular cytogenetic landmarks. These new genomic resources allowed refinement of the syntenic relationships among C. hystrix, cucumber and melon (C. melo, 2n = 2x = 24) chromosomes. An improved model was proposed to elucidate the evolutionary history in which the seven cucumber chromosomes were evolved from an x=12 ancestor through dysploid chromosome reduction that involved in four translocations, four chromosomal fusions, and 53 inversions. A synthetic tetraploid, C. x hytivus (2n = 4x = 38, HHCC) was developed through induction of chromosome doubling of the interspecific hybrid between C. sativus and C. hystrix, which was used as the bridge to develop an introgression library through marker-assisted backcrossing. The molecularly characterized introgression lines provide insights into homoeologous pairing between cucumber and C. hystrix chromosomes. Supported by Agriculture and Food Research Initiative Competitive Grant nos. 2013-67013-21105 and 2015- 51181-24285. Partial Funding provided by the Specialty Crop Block Grant Program at the U.S. Department of Agriculture (USDA) through Grant 14-SCBGP-CA-0006 (The CucCAP Project)
The USDA Cucumber (Cucumis sativus L.) Collection: Genetic Diversity, Population Structure, Genome-Wide Association Studies and Core Collection Development
Xin Wang1, Kan Bao1, Umesh K.Reddy2, Yang Bai1, Sue A. Hammer3, Chen Jiao1, Todd C. Wehner4, Axel O. Ramírez-Madera5, Yiqun Weng5, Rebecca Grumet3, Zhangjun Fei1
1Boyce Thompson Institute, Ithaca, NY, USA. 2Gus R. Douglass Institute and Department of Biology, West Virginia State University, Institute, WV, WV, USA. 3Department of Horticulture, Michigan State University, East Lansing, MI, USA. 4Horticultural Science Department, North Carolina State University, Raleigh, NC, USA. 5U.S. Department of Agriculture, Agricultural Research Service, Vegetable Crops Research Unit, Madison, WI, USA
Germplasm collections are a crucial resource to conserve natural genetic diversity and provide a source of novel traits essential for sustained crop improvement. Optimal collection, preservation and utilization of these materials depends upon knowledge of the genetic variation present within the collection. Here we use the high throughput genotyping-by- sequencing (GBS) technology to characterize the United States National Plant Germplasm System (NPGS) collection of cucumber (Cucumis sativus L.). The GBS data, derived from 1,234 cucumber accessions, provided more than 23K high quality single nucleotide polymorphisms (SNPs) that are well distributed at high density in the genome (~1 SNP/10.6 kb). The SNP markers were used to characterize genetic diversity, population structure, phylogenetic relationships, linkage disequilibrium, and population differentiation of the NPGS cucumber collection. These results, providing detailed genetic analysis of the U.S. cucumber collection, complement NPGS descriptive information regarding geographic origin and phenotypic characterization. We also identified genome regions significantly associated with thirteen horticulturally important traits through genome-wide association studies (GWAS). Finally, we developed a molecularly-informed, publicly accessible core collection of 395 accessions that represents at least 96% of the genetic variation present in the NPGS. Collectively, the information obtained from the GBS data enabled deep insight into the diversity present and genetic relationships among accessions within the collection, and will provide a valuable resource for genetic analysis, gene discovery, crop improvement and germplasm preservation. Partial Funding provided by the Specialty Crop Block Grant Program at the U.S. Department of Agriculture (USDA) through Grant 14-SCBGP-CA-0006 (The CucCAP Project)
Genome-Wide Identification of Calcium-Dependent Protein Kinase and its Related Kinase Gene Families in Cucurbitaceae Species
Chunhua Wei, Xian Zhang
Northwest A&F university, Yangling, Shaanxi, China
Both the calcium-dependent protein kinase (CDPK) and CDPK-related kinases (CRKs) play numerous roles related to plant growth, development, and stress response. Despite genome- wide identification of both families in Cucumis species, comparative evolutionary and functional analysis of both CDPKs and CRKs in Cucurbitaceae remain unclear. Here, we totally identified 128 CDPK and 56 CRK genes in six Cucurbitaceae species (C. lannatus, C. sativus, C. moschata, C. maxima, C. pepo, and L. siceraria). Structural variation analysis indicated that two CDPKs (CpCDPK19 and CpCDPK27) in C. pepo may have undergone subfunctionalization via self- duplication of conserved domains. Using the watermelon genome as reference, an integrated map containing 23 loci (16 CDPK and nine CRK loci) were obtained, 16 of which (12 CDPK and four CRK) were shared by all seven Cucurbitaceae species. Phylogenetic analysis obtained four CDPK groups and one CRK group with one or two major intron phase patterns. Moreover, the topologies of most loci are consistent with a recently published evolutionary scenario of seven modern Cucurbitaceae species. Comparative syntenic analysis detected few segmental duplication events in Benincaseae tribe species, but many in Cucurbita tribe species. In addition, expression patterns of ClCDPKs and ClCRKs were studied under different abiotic stresses, as well as subcellular localizations of several ClCDPKs and ClCRKs.
Transcriptome Dynamics of the Whitefly Bemisia tabaci in Response to Feeding on Melon Plants Infected with Cucurbit yellow stunting disorder virus
Navneet Kaur1, Wenbo Chen², Zhangjun Fei², William M. Wintermantel¹
¹U.S. Department of Agriculture, Agricultural Research Service, Crop Improvement and Protection Research Unit, Salinas, CA, USA. ²Boyce Thompson Institute, Ithaca, NY, USA
Whiteflies are a serious threat to crop production. They can cause direct damage through feeding, as well as indirect damage through production of honeydew leading to sooty molds, and particularly through transmission of plant viruses. Whiteflies are known to transmit economically important viruses in the genera Begomovirus, Crinivirus, Ipomovirus, Carlavirus, and Torradovirus, and all are known to cause economic loss to the production of cucurbit crops. Recent studies have demonstrated common patterns of gene regulation between whiteflies fed on tomato infected with either a crinivirus or begomovirus (Kaur et al. 2017; BMC Genomics, 18: 370; Hasegawa et al., 2018, Virology 513: 52). In order to gain a better understanding of the whitefly response to virus-infected cucurbits and cucurbit viruses, RNA-Seq was performed on whiteflies following acquisition feeding on melon leaves infected with the semipersistently transmitted crinivirus, Cucurbit yellow stunting disorder virus (CYSDV) for three different time periods, 24 h, 72 h, and 7 days. CYSDV is transmitted exclusively by the whitefly, Bemisia tabaci, and can be retained for seven to nine days in the vector. A total of 275 differentially expressed genes (DEGs) were identified in response to feeding on CYSDV-infected melon plants. Interestingly, only 3 DEGs (all down-regulated) were observed at 24 h, followed by 221 DEGs at 72 h, and 51 DEGs at 7 days. Several distinct gene categories were represented among the DEGs in the whiteflies. As was found in previous studies involving another crinivirus, Tomato chlorosis virus (ToCV), a large percentage of the DEGs were orphan genes that are unique to the whitefly and do not show any homology to known genes in other species. Further, we found 59 DEGs common between whiteflies fed on CYSDV-infected melon and ToCV-infected tomato plants, and 14 in common between those fed on CYSDV, ToCV and the begomovirus, Tomato yellow leaf curl virus, suggesting certain common responses by whiteflies to feeding on crinivirus- infected host plants and perhaps virus-infected plants in general, that can be harnessed to interfere with transmission to melon and other crops.
Pan-Genomes of Citrullus Species
Shan Wu1, Lei Gao1, John Mendieta1, Shaogui Guo2, Honghe Sun2, Yong Xu2, Shaker Kousik3, W. Patrick Wechter3, Amnon Levi3, Zhangjun Fei1
1Boyce Thompson Institute, Ithaca, NY, USA. 2National Engineering Research Center for Vegetables, Beijing, China. 3U.S. Department of Agriculture, Agricultural Research Service, U.S. Vegetable Laboratory, Charleston, SC, USA
Watermelon (Citrullus lanatus) is among the most important vegetable crops in the world. It belongs to the Citrullus (2n=2x=22) genus, which includes six other species, namely egusi watermelon (C. mucosospermus), citron watermelon (C. amarus), colocynth (C. colocynthis), C. ecirrhosus, C. rehmii and C. naudinianus. To maximize the capture of genome variations within and among these Citrullus species and to identify novel agronomically important alleles for facilitating watermelon breeding, we are constructing pan-genomes of the Citrullus species. We have been generating high-quality reference genomes from selected individuals of four Citrullus species, including two C. lanatus accessions, two C. amarus and one C. colocynthis whose genome assemblies and annotations are finished, and one in the process for C. mucosospermus. We also generated whole genome resequencing data from more than 400 additional accessions of these four species, which are being de novo assembled for each accession. A pan-genome for each of the four species will be constructed by combining the reference genome(s) and de novo assembled novel non-redundant sequences. Presence-absence variations (PAVs) of protein- coding genes will be analyzed, and collections of core and distributed gene sets for each species will be identified. Comparative analysis of the four pan-genomes will be performed to highlight syntenic regions and species-specific variations. The Citrullus pan-genomes and the identified PAVs will enable us to trace the distribution of functionally important alleles and gain insights into watermelon evolution, domestication and introgression, and provide an important resource to facilitate the mining of natural variation in Citrullus for scientific studies and the improvement of watermelon. Partial Funding provided by the Specialty Crop Block Grant Program at the U.S. Department of Agriculture (USDA) through Grant 14-SCBGP-CA-0006 (The CucCAP Project)
Developing Genome and transcriptome Information Database for functional Genomics Research of Japanese Muskmelon
Ryoichi Yano1,2,3, Tohru Ariizumi1,4, Satoko Nonaka1,4, Hiroshi Ezura4,1
1Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Japan. 2Advanced Analysis Center, National Agriculture and Food Research Organization, Tsukuba, Japan. 3JST PRESTO, Kawaguchi, Japan. 4Tsukuba-Plant Innovation Research Center, University of Tsukuba, Tsukuba, Japan
Melon (Cucumis melo L.) is an important Cucurbitaceae crop that is widely produced in the world. It exhibits a wide range of natural variation especially in fruit phenotypes (i.e., climacteric and non-climacteric ripening types). The semi-climacteric type inbred cultivar ‘Earl’s favorite Harukei-3’ (Harukei-3) is known for its sweetness and rich aroma, and hence popular as a breeding material for development of high-grade muskmelon in Japan. To promote functional genomics research and breeding of Japanese muskmelon, we have developed gene information database ‘Melonet-DB’ (http://gene.melonet-db.jp). In the database, a user is able to visually compare tissue-specific gene expression patterns as well as gene coexpression across multiple input queries. To update Melonet-DB, we recently conducted additional RNA-seq studies, and the updated dataset will contain new RNA-seq datasets such as imbibed seeds, seedlings, and post-harvest ethylene-emitting fruit tissues. In addition, by combining a non-destructive fruit phenotype analysis method, leaf RNA-seq datasets have been also collected in a weekly manner in a greenhouse to analyze sink-source tissue interaction. In addition to the transcriptome study, the whole genome information of the Harukei-3 melon has been also developed using the 2nd and 3rd next-generation DNA sequencers. By combining 43-fold Pacbio long reads, 190- fold Bionano single molecule data, 82-fold Illumina Hiseq short reads, and 52-fold mate pair reads, we assembled Harukei-3 genomic scaffolds. Some scaffolds span chromosome arm, indicating that combining these technologies was effective to construct long scaffolds. In this presentation, we’d like to introduce the results of our recent research activities.