Melissa H. Jia

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.

RL-SAGE and microarray analysis of the rice transcriptome after Rhizoctonia solani infection

Sheath blight caused by the fungal pathogen Rhizoctonia solani is an emerging problem in rice production worldwide. To elucidate the molecular basis of rice defense to the pathogen, RNA isolated from R. solani-infected leaves of Jasmine 85 was used for both RL-SAGE library construction and microarray hybridization. RL-SAGE sequence analysis identified 20,233 and 24,049 distinct tags from the control and inoculated libraries, respectively. Nearly half of the significant tags (≥2 copies) from both libraries matched TIGR annotated genes and KOME full-length cDNAs. Among them, 42% represented sense and 7% antisense transcripts, respectively. Interestingly, 60% of the library-specific (≥10 copies) and differentially expressed (>4.0-fold change) tags were novel transcripts matching genomic sequence but not annotated genes. About 70% of the genes identified in the SAGE libraries showed similar expression patterns (up or down-regulated) in the microarray data obtained from three biological replications. Some candidate RL-SAGE tags and microarray genes were located in known sheath blight QTL regions. The expression of ten differentially expressed RL-SAGE tags was confirmed with RT-PCR. The defense genes associated with resistance to R. solani identified in this study are useful genomic materials for further elucidation of the molecular basis of the defense response to R. solani and fine mapping of target sheath blight QTLs.

Factors Affecting the Outcrossing Rate between Clearfield™ Rice and Red Rice (Oryza sativa)

Abstract The commercialization of imazethapyr-resistant (Clearfield™, CL) rice in the southern United States has raised serious concerns about gene flow to red rice, producing imazethapyr-resistant red rice populations. Our objectives were to determine the impact of planting date, CL cultivars, and red rice biotypes on outcrossing rate; and to investigate the relative contribution of flowering time of CL rice and red rice biotypes, together with air temperature and relative humidity (RH), on outcrossing rate. Field experiments were conducted at Stuttgart, Rohwer, and Kibler, AR, from 2005 to 2007, at three or four planting times from mid-April to late May. ‘CL161’ (inbred cultivar) and ‘CLXL8’ (hybrid) rice were planted in nine-row plots, with red rice planted in the middle row. Twelve red rice biotypes were used. The flowering of red rice and CL rice, air temperature, and RH were recorded. Red rice seeds were collected at maturity. To estimate outcrossing rate, resistance to imazethapyr was evaluated in subsequent years and confirmed using rice microsatellite markers. CLXL8 rice flowered 2 to 4 d earlier than CL161 rice, and flowering was completed within 1 wk in all plantings. The flowering duration of most red rice biotypes ranged from 4 to 17 d. Flowering synchrony of red rice biotypes and CL rice ranged from 0 to 100% at different plantings. In general, CLXL8 had greater flowering overlap and higher outcrossing rate with red rice than did CL161 rice. The outcrossing rate of red rice biotypes ranged from 0 to 0.21% and 0 to 1.26% with CL161 and CLXL8 rice, respectively. The outcrossing rate differed within each planting date (P < 0.05). Outcrossing was generally lower in mid-May and late May than in mid-April and late April planting times. Flowering synchrony and outcrossing rate were not correlated (r2 < 0.01). Outcrossing with CL161 was primarily influenced by red rice biotype. A minimum air temperature of > 24 C in the evening also favors outcrossing with CL161. With CLXL8 rice, outcrossing was most affected by RH. When RH was < 54%, outcrossing was less (0.12%) than when RH was ≥ 54% (0.38%). With CLXL8 rice, a minimum RH of ≥ 54%, from mid-morning to noon, increased outcrossing with red rice. To fully understand the interaction effects of these factors on outcrossing with red rice, controlled experiments are needed. Nomenclature: Imazethapyr; Red rice, Oryza sativa L.; Rice, Oryza sativa L. ORYSA.

Similar traits, different genes? Examining convergent evolution in related weedy rice populations

Convergent phenotypic evolution may or may not be associated with convergent genotypic evolution. Agricultural weeds have repeatedly been selected for weed‐adaptive traits such as rapid growth, increased seed dispersal and dormancy, thus providing an ideal system for the study of convergent evolution. Here, we identify QTL underlying weedy traits and compare their genetic architecture to assess the potential for convergent genetic evolution in two distinct populations of weedy rice. F2 offspring from crosses between an indica cultivar and two individuals from genetically differentiated U.S. weedy rice populations were used to map QTL for four quantitative (heading date, seed shattering, plant height and growth rate) and two qualitative traits. We identified QTL on nine of the twelve rice chromosomes, yet most QTL locations do not overlap between the two populations. Shared QTL among weed groups were only seen for heading date, a trait for which weedy groups have diverged from their cultivated ancestors and from each other. Sharing of some QTL with wild rice also suggests a possible role in weed evolution for genes under selection during domestication. The lack of overlapping QTL for the remaining traits suggests that, despite a close evolutionary relationship, weedy rice groups have adapted to the same agricultural environment through different genetic mechanisms.

Molecular Evolution of the Rice Blast Resistance Gene Pi-ta in Invasive Weedy Rice in the USA

The Pi-ta gene in rice has been effectively used to control rice blast disease caused by Magnaporthe oryzae worldwide. Despite a number of studies that reported the Pi-ta gene in domesticated rice and wild species, little is known about how the Pi-ta gene has evolved in US weedy rice, a major weed of rice. To investigate the genome organization of the Pi-ta gene in weedy rice and its relationship to gene flow between cultivated and weedy rice in the US, we analyzed nucleotide sequence variation at the Pi-ta gene and its surrounding 2 Mb region in 156 weedy, domesticated and wild rice relatives. We found that the region at and around the Pi-ta gene shows very low genetic diversity in US weedy rice. The patterns of molecular diversity in weeds are more similar to cultivated rice (indica and aus), which have never been cultivated in the US, rather than the wild rice species, Oryza rufipogon. In addition, the resistant Pi-ta allele (Pi-ta) found in the majority of US weedy rice belongs to the weedy group strawhull awnless (SH), suggesting a single source of origin for Pi-ta. Weeds with Pi-ta were resistant to two M. oryzae races, IC17 and IB49, except for three accessions, suggesting that component(s) required for the Pi-ta mediated resistance may be missing in these accessions. Signatures of flanking sequences of the Pi-ta gene and SSR markers on chromosome 12 suggest that the susceptible pi-ta allele (pi-ta), not Pi-ta, has been introgressed from cultivated to weedy rice by out-crossing.

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.

Identification of two major resistance genes against race IE-1k of Magnaporthe oryzae in the indica rice cultivar Zhe733

The race IE-1k of Magnaporthe oryzae recovered from the Southern US overcomes the resistance (R) gene Pita. The objectives of the present study were to identify and tag R genes to IE-1k for rice breeding. TM2, S1, 94071, and B isolates of the race IE-1k were used to identify and map R genes from a resistant indica rice cultivar Zhe733 using a recombinant inbred line population from a cross of the genetic stock KBNTlpa1-1 and Zhe733. The ratio of 3 resistant:1 susceptible in 162 RIL of an F10-11 KBNTlpa1-1/Zhe733 (K/Z) population indicated that two major R genes in Zhe733 confer resistance to IE-1k. A total of 118 polymorphic simple sequence repeat markers were analyzed in 162 F10-11 individuals of the K/Z population to determine chromosomal locations of the loci conferring resistance to race IE-1k using composite interval mapping. Two major R genes temporarily designated as Pi42(t) and Pi43(t) each providing complete resistance to IE-1k were identified on chromosomes 8 and 11, respectively. RILs containing Pi42(t) and Pi43(t) were also resistant to other US races IB-1, IB-45, IB-49, IB-54, IC-17, IE-1, IG-1, and IH-1. The Pi42(t) gene was mapped between RM310 and RM72, and the location of Pi43(t) was closely associated with two flanking SSR markers RM1233 and RM224 on chromosome 11 in a chromosomal region carrying the resistance gene Pi1. Two molecular markers RM72 and RM1233 identified in this study should be useful for fine mapping and for facilitating incorporation of Pi42(t) and Pi43(t) into advanced breeding lines by marker-assisted selection.

Genome-Wide Association of Rice Blast Disease Resistance and Yield-Related Components of Rice.

Robust disease resistance may require an expenditure of energy that may limit crop yield potential. In the present study, a subset of a United States Department of Agriculture rice core collection consisting of 151 accessions was selected using a major blast resistance (R) gene, Pi-ta, marker and was genotyped with 156 simple sequence repeat (SSR) markers. Disease reactions to Magnaporthe oryzae, the causal agent of rice blast disease, were evaluated under greenhouse and field conditions, and heading date, plant height, paddy and brown seed weight in two field environments were analyzed, using an association mapping approach. A total of 21 SSR markers distributed among rice chromosomes 2 to 12 were associated with blast resistance, and 16 SSR markers were associated with seed weight, heading date, and plant height. Most noticeably, shorter plants were significantly correlated with resistance to blast, rice genomes with Pi-ta were associated with lighter seed weights, and the susceptible alleles of RM171 and RM6544 were associated with heavier seed weight. These findings unraveled a complex relationship between disease resistance and yield-related components.

Identification of the Rice Blast Resistance Gene Pib in the National Small Grains Collection

The Pib gene in rice confers resistance to a wide range of races of the rice blast pathogen, Magnaporthe oryzae, including race IE1k that overcomes Pita, another broad-spectrum resistance gene. In this study, the presence of Pib was determined in 164 rice germplasm accessions from a core subset of the National Small Grains Collection utilizing DNA markers and pathogenicity assays. The presence of Pib was evaluated with two simple sequence repeat (SSR) markers and a dominant marker (Pib-dom) derived from the Pib gene sequence. Pathogenicity assays using two avirulent races (IE1k and IB1) and a virulent race (IB54) were performed to verify the resistance responses of accessions. Of the 164 accessions evaluated, 109 contained the Pib gene as determined using both SSR markers and pathogenicity assays, albeit different haplotypes were detected. The remaining 52 germplasm accessions were different in their responses to the blast races IB54, IE1k, and IB1, thus indicating the presence of R gene(s) other than Pib. The accessions characterized in this study could be used for marker-assisted breeding to improve blast resistance in indica and japonica cultivars worldwide.

Field performance of STG06L-35-061, a new genetic resource developed from crosses between weed-suppressive indica rice and commercial southern U.S. long-grains

AimsWeed control in rice is challenging, particularly in light of increased resistance to herbicides in weed populations including Echinochloa crus-galli (L.) Beauv. Indica rice cultivars can produce high yields and suppress barnyardgrass, but have not been commercially acceptable in the U.S. due to inferior agronomic traits and grain quality. Our objectives were to combine high yield and weed-suppressive characteristics from indica cultivars with commercially acceptable grain quality and plant types from long-grain cultivars grown in the southern U.S.MethodsCrosses between indica and commercial tropical japonica (cv. Katy, and cv. Drew) rice were evaluated for weed suppression and agronomic traits in a breeding program.ResultsIn some tests, the selection STG06L-35-061 was nearly as weed suppressive as PI 312777, the suppressive parent, and more suppressive than its tropical japonica parents. Its main crop yield is commercially acceptable, and intermediate between PI 312777 and Katy. Its milling quality and cooking quality are similar to long-grain commercial cultivars, and it has resistance to rice blast disease. Marker analyses identified introgressions from the indica parents on chromosomes 1 and 3 of STG06L-35-061 that require further analysis as possible sources of weed suppressive traits.ConclusionsSTG06L-35-061 might be suitable for organic rice or reduced input conventional systems.