Aaron Jackson

Genotypic and phenotypic characterization of genetic differentiation and diversity in the USDA rice mini-core collection

A rice mini-core collection consisting of 217 accessions has been developed to represent the USDA core and whole collections that include 1,794 and 18,709 accessions, respectively. To improve the efficiency of mining valuable genes and broadening the genetic diversity in breeding, genetic structure and diversity were analyzed using both genotypic (128 molecular markers) and phenotypic (14 numerical traits) data. This mini-core had 13.5 alleles per locus, which is the most among the reported germplasm collections of rice. Similarly, polymorphic information content (PIC) value was 0.71 in the mini-core which is the highest with one exception. The high genetic diversity in the mini-core suggests there is a good possibility of mining genes of interest and selecting parents which will improve food production and quality. A model-based clustering analysis resulted in lowland rice including three groups, aus (39 accessions), indica (71) and their admixtures (5), upland rice including temperate japonica (32), tropical japonica (40), aromatic (6) and their admixtures (12) and wild rice (12) including glaberrima and four other species of Oryza. Group differentiation was analyzed using both genotypic distance Fst from 128 molecular markers and phenotypic (Mahalanobis) distance D2 from 14 traits. Both dendrograms built by Fst and D2 reached similar-differentiative relationship among these genetic groups, and the correlation coefficient showed high value 0.85 between Fst matrix and D2 matrix. The information of genetic and phenotypic differentiation could be helpful for the association mapping of genes of interest. Analysis of genotypic and phenotypic diversity based on genetic structure would facilitate parent selection for broadening genetic base of modern rice cultivars via breeding effort.

A natural mutation in rc reverts white-rice-pericarp to red and results in a new, dominant, wild-type allele: Rc-g

The Rc locus regulates pigmentation of the rice bran layer, and selection for the rc allele (white pericarp) occurred during domestication of the crop. White bran is now ubiquitous among cultivated varieties throughout rice growing regions of the world. We identified a new allele that arose by natural mutation within the rc pseudogene of the cultivar ‘Wells’. The mutation restored the reading frame of the gene, and reverted the bran layer pigmentation to red (wild-type). By sequencing the Rc locus in plants derived from red seeds, and linkage analysis in a segregating population, we were able to demonstrate that mutation within rc resulted in the new, dominant, wild-type allele Rc-g.

Analysis of rice blast resistance gene Pi-z in rice germplasm using pathogenicity assays and DNA markers

The Pi-z gene in rice confers resistance to a wide range of races of the rice blast fungus, Magnaporthe oryzae. The objective of this study was to characterize Pi-z in 111 rice germplasm accessions using DNA markers and pathogenicity assays. The existence of Pi-z in rice germplasm was detected by using four simple sequence repeat (SSR) markers (RM527, AP4791, AP5659-1, AP5659-5) closely linked to Pi-z, and was verified using pathogenicity assays with an avirulent strain (IE1k) and two virulent races (IB33 and IB49). Among 111 germplasm accessions evaluated, 73 were found to contain the Pi-z gene using both SSR markers and pathogenicity assays. The remaining 38 germplasm accessions were found to be inconsistent in their responses to the blast races IB33, IEIk and IB49 with expected SSR marker alleles, suggesting the presence of unexpected SSR alleles and additional R gene(s). These characterized germplasm can be used for genetic studies and marker-assisted breeding for improving blast resistance in rice.

Characterization and comparative profiling of the small RNA transcriptomes in two phases of flowering in Cymbidium ensifolium

BackgroundCymbidium ensifolium is one of the most important ornamental flowers in China, with an elegant shape, beautiful appearance, and a fragrant aroma. Its unique flower shape has long attracted scientists. MicroRNAs (miRNAs) are critical regulators in plant development and physiology, including floral development. However, to date, few studies have examined miRNAs in C. ensifolium.ResultsIn this study, we employed Solexa technology to sequence four small RNA libraries from two flowering phases to identify miRNAs related to floral development. We identified 48 mature conserved miRNA and 71 precursors. These conserved miRNA belonged to 20 families. We also identified 45 novel miRNA which includes 21 putative novel miRNAs*, and 28 hairpin forming precursors. Two trans-acting small interfering RNAs (ta-siRNAs) were identified, one of which was homologous to TAS3a1. TAS3a1 belongs to the TAS3 family, which has been previously reported to target auxin response factors (ARF) and be involved in plant growth and floral development. Moreover, we built a C. ensifolium transctriptome database to identify genes targeted by miRNA, which resulted in 790 transcriptomic target unigenes. The target unigenes were annotated with information from the non-redundant (Nr), gene ontology database (GO), eukaryotic orthologous groups (KOGs) and Kyoto encyclopedia of genes and genomes (KEGG) database. The unigenes included MADS-box transcription factors targeted by miR156, miR172 and miR5179, and various hormone responding factors targeted by miR159. The MADS-box transcription factors are well known to determine the identity of flower organs and hormone responding factors involved in floral development. In expression analysis, three novel and four conserved miRNA were differentially expressed between two phases of flowering. The results were confirmed by RNA-seq and qRT-PCR. The differential expression of two miRNA, miR160 and miR396, targeted ARFs and growth regulating factor (GRF), respectively. However, most of these small RNA were clustered in the uncharacterized group, which suggests there may be many novel small non-coding RNAs yet to be discovered.ConclusionOur study provides a diverse set of miRNAs related to cymbidium floral development and serves as a useful resource for investigating miRNA-mediated regulatory mechanisms of floral development.

Development of Cymbidium ensifolium genic-SSR markers and their utility in genetic diversity and population structure analysis in cymbidiums.

BackgroundCymbidium is a genus of 68 species in the orchid family, with extremely high ornamental value. Marker-assisted selection has proven to be an effective strategy in accelerating plant breeding for many plant species. Analysis of cymbidiums genetic background by molecular markers can be of great value in assisting parental selection and breeding strategy design, however, in plants such as cymbidiums limited genomic resources exist. In order to obtain efficient markers, we deep sequenced the C. ensifolium transcriptome to identify simple sequence repeats derived from gene regions (genic-SSR).ResultThe 7,936 genic-SSR markers were identified. A total of 80 genic-SSRs were selected, and primers were designed according to their flanking sequences. Of the 80 genic-SSR primer sets, 62 were amplified in C. ensifolium successfully, and 55 showed polymorphism when cross-tested among 9 Cymbidium species comprising 59 accessions. Unigenes containing the 62 genic-SSRs were searched against Non-redundant (Nr), Gene Ontology database (GO), eukaryotic orthologous groups (KOGs) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database. The search resulted in 53 matching Nr sequences, of which 39 had GO terms, 18 were assigned to KOGs, and 15 were annotated with KEGG. Genetic diversity and population structure were analyzed based on 55 polymorphic genic-SSR data among 59 accessions. The genetic distance averaged 0.3911, ranging from 0.016 to 0.618. The polymorphic index content (PIC) of 55 polymorphic markers averaged 0.407, ranging from 0.033 to 0.863. A model-based clustering analysis revealed that five genetic groups existed in the collection. Accessions from the same species were typically grouped together; however, C. goeringii accessions did not always form a separate cluster, suggesting that C. goeringii accessions were polyphyletic.ConclusionThe genic-SSR identified in this study constitute a set of markers that can be applied across multiple Cymbidium species and used for the evaluation of genetic relationships as well as qualitative and quantitative trait mapping studies. Genic-SSR’s coupled with the functional annotations provided by the unigenes will aid in mapping candidate genes of specific function.

Genetic Variation and Association Mapping of Protein Concentration in Brown Rice Using a Diverse Rice Germplasm Collection

ABSTRACT Protein is the second most abundant constituent in the rice grain next to starch. Association analysis for protein concentration in brown rice was performed using a “mini-core” collection, which represents the germplasm diversity found in the USDA rice world collection. Protein concentration was determined in replicated trials conducted in two southern U.S. locations, and association mapping was performed by using 157 genomewide DNA markers. Protein concentration ranged from 5.4 to 11.9% among the 202 accessions. Protein variation owing to accession and accession × location interaction were highly significant. Ample variation was seen within each subpopulation by ancestry, as well as within the 14 geographic regions where the accessions originated. Accessions from Eastern Europe had the highest level of protein. Ten markers on eight chromosomes were significantly associated with protein concentration. Five of these markers occurred near known protein precursor genes or quantitative trait loci, an...

Outcrossing Potential between U.S. Blackhull Red Rice and Indica Rice Cultivars

Abstract Weedy red rice is a major weed pest of rice in the southern United States. Outcrossing between red rice and commercial tropical japonica rice cultivars has resulted in new weed biotypes that further hinder the effectiveness of weed management. In recent years, indica rice has been used increasingly as a germplasm source for breeding and for reduced-input systems in the United States, but little is known about its outcrossing potential with U.S. weedy red rice biotypes. In a 2-yr study, simple sequence repeat marker analysis was used to show that blackhull (BH) red rice (PI 653424) outcrossing to four, late-maturing indica cultivars averaged 0.0086% and ranged from 0.002% for ‘TeQing’ to 0.0173% for ‘4484’ (PI 615022). Rates of outcrossing to a tropical japonica cultivar standard, ‘Kaybonnet’ (0.032%), were substantially greater than for the indica cultivars. These differences in outcrossing were due largely to synchronization of flowering times between rice and red rice, with Kaybonnet and TeQing exhibiting the greatest and least synchronization, respectively. Outcrossing rates also may have been affected by rice–red rice flower density differences within the rice plots. Outcrossing from cultivated rice to the red rice (as pollen recipient), which was taller than all rice cultivars, was undetectable in these studies, and environmental conditions (e.g., temperature, humidity, solar radiation, and rainfall) were not strongly correlated with the outcrossing rates observed. Grain yields of the original BH red rice line were greatest in the Kaybonnet plots, demonstrating that the indica cultivars were superior competitors against this weed. Collectively, these results suggest that red rice biotypes that flower synchronously with rice cultivars are a potential source of pollen for outcrossing and gene flow in rice fields in the southern United States. Nomenclature: Red rice, Oryza sativa L.; rice, Oryza sativa L.; indica rice cultivars ‘TeQing’, ‘4484’; tropical japonica rice cultivar ‘Kaybonnet’.

Development of genetic markers linked to straighthead resistance through fine mapping in rice (Oryza sativa L.).

Straighthead, a physiological disorder characterized by sterile florets and distorted spikelets, causes significant yield losses in rice, and occurs in many countries. The current control method of draining paddies early in the season stresses plants, is costly, and wastes water. Development of resistant cultivar is regarded as the most efficient way for its control. We mapped a QTL for straighthead resistance using two recombinant inbred line (RIL) F9 populations that were phenotyped over two years using monosodium methanearsonate (MSMA) to induce the symptoms. One population of 170 RILs was genotyped with 136 SSRs and the other population of 91 RILs was genotyped with 159 SSRs. A major QTL qSH-8 was identified in an overlapping region in both populations, and explained 46% of total variation in one and 67% in another population for straighthead resistance. qSH-8 was fine mapped from 1.0 Mbp to 340 kb using 7 SSR markers and further mapped to 290 kb in a population between RM22573 and InDel 27 using 4 InDel markers. SSR AP3858-1 and InDel 11 were within the fine mapped region, and co-segregated with straighthead resistance in both RIL populations, as well as in a collection of diverse global accessions. These results demonstrate that AP3858-1 and InDel 11 can be used for marker-assisted selection (MAS) for straighthead resistant cultivars, which is especially important because there is no effective way to directly evaluate straighthead resistance.

Erratum to: Genetic analysis of genetic basis of a physiological disorder “straighthead” in rice (Oryza sativa L.)

Unfortunately, the original version of this article contains a few errors. Figure 2 in the original publication, qSTH-2 and qSTH-8 is quite displaced, and the arrow for qSTH-8 is not shown. The corrected figure is shown in this erratum. Moreover, the third author, Hesham Agrama, has added another affiliation. His additional affiliation is: Department of Crop Sciences, College of Agricultural and Marine Sciences Sultan Qaboos University P.O. Box 34, Alkhoud 123, Oman.