Microbiome properties in the root nodules of Prosopis cineraria, a leguminous desert tree.

We conducted a comprehensive analysis of the total microbiome and transcriptionally active microbiome communities in the roots and root nodules of Prosopis cineraria, an important leguminous tree in arid regions of many Asian countries. Mature P. cineraria trees growing in the desert did not exhibit any detected root nodules. However, we observed root nodules on the roots of P. cineraria growing on a desert farm and on young plants growing in a growth chamber, when inoculated with rhizosphere soil, including with rhizosphere soil from near desert tree roots that had no nodules. Compared to nearby soil, non-nodulated roots were enriched with Actinobacteria (e.g., Actinophytocola sp.), whereas root nodules sampled from the desert farm and growth chamber had abundant Alphaproteobacteria (e.g., Ensifer sp.). These nodules yielded many microbes in addition to such nitrogen-fixing bacteria as Ensifer and Sinorhizobium species. Significant differences exist in the composition and abundance of microbial isolates between the nodule surface and the nodule endosphere. Shotgun metagenome analysis of nodule endospheres revealed that the root nodules comprised over 90% bacterial DNA, whereas metatranscriptome analysis showed that the plant produces vastly more transcripts than the microbes in these nodules. Control inoculations demonstrated that four out of six Rhizobium, Agrobacterium, or Ensifer isolates purified from P. cineraria nodules produced nodules in the roots of P. cineraria seedlings under greenhouse conditions. The best nodulation was achieved when seedlings were inoculated with a mixture of those bacterial strains. Though root nodulation could be achieved under water stress conditions, nodule number and nodule biomass increased with copious water availability. .IMPORTANCEMicrobial communities were investigated in roots and root nodules of Prosopis cineraria, a leguminous tree species in arid Asian regions that is responsible for exceptionally important contributions to soil fertility in these dramatically dry locations. Soil removed from regions near nodule-free roots on these mature plants contained an abundance of bacteria with the genetic ability to generate nodules and fix nitrogen but did not normally nodulate in their native rhizosphere environment, suggesting a very different co-evolved relationship than that observed for herbaceous legumes. The relative over-expression of the low-gene-density plant DNA compared to the bacterial DNA in the nodules was also unexpected, indicating a very powerful induction of host genetic contributions within the nodule. Finally, the water dependence of nodulation in inoculated seedlings suggested a possible link between early seedling growth (before a deep root system can be developed) and the early development of nitrogen-fixing capability.

Identification of microRNAs responsive to arbuscular mycorrhizal fungi in Panicum virgatum (switchgrass).

BACKGROUND: MicroRNAs (miRNAs) are important post-transcriptional regulators involved in the control of a range of processes, including symbiotic interactions in plants. MiRNA involvement in arbuscular mycorrhizae (AM) symbiosis has been mainly studied in model species, and our study is the first to analyze global miRNA expression in the roots of AM colonized switchgrass (Panicum virgatum), an emerging biofuel feedstock. AM symbiosis helps plants gain mineral nutrition from the soil and may enhance switchgrass biomass production on marginal lands. Our goals were to identify miRNAs and their corresponding target genes that are controlling AM symbiosis in switchgrass. RESULTS: Through genome-wide analysis of next-generation miRNA sequencing reads generated from switchgrass roots, we identified 122 mature miRNAs, including 28 novel miRNAs. By comparing miRNA expression profiles of AM-inoculated and control switchgrass roots, we identified 15 AM-responsive miRNAs across lowland accession "Alamo", upland accession "Dacotah", and two upland/lowland F1 hybrids. We used degradome sequencing to identify target genes of the AM-responsive miRNAs revealing targets of miRNAs residing on both K and N subgenomes. Notably, genes involved in copper ion binding were targeted by downregulated miRNAs, while upregulated miRNAs mainly targeted GRAS family transcription factors. CONCLUSION: Through miRNA analysis and degradome sequencing, we revealed that both upland and lowland switchgrass genotypes as well as upland-lowland hybrids respond to AM by altering miRNA expression. We demonstrated complex GRAS transcription factor regulation by the miR171 family, with some miR171 family members being AM responsive while others remained static. Copper miRNA downregulation was common amongst the genotypes tested and we identified superoxide dismutases and laccases as targets, suggesting that these Cu-miRNAs are likely involved in ROS detoxification and lignin deposition, respectively. Other prominent targets of the Cu miRNAs were blue copper proteins. Overall, the potential effect of AM colonization on lignin deposition pathways in this biofuel crop highlights the importance of considering AM and miRNA in future biofuel crop development strategies.

The Genome of the Mimosoid Legume Prosopis cineraria, a Desert Tree.

The mimosoid legumes are a clade of ~40 genera in the Caesalpinioideae subfamily of the Fabaceae that grow in tropical and subtropical regions. Unlike the better studied Papilionoideae, there are few genomic resources within this legume group. The tree Prosopis cineraria is native to the Near East and Indian subcontinent, where it thrives in very hot desert environments. To develop a tool to better understand desert plant adaptation mechanisms, we sequenced the P. cineraria genome to near-chromosomal assembly, with a total sequence length of ~691 Mb. We predicted 77,579 gene models (76,554 CDS, 361 rRNAs and 664 tRNAs) from the assembled genome, among them 55,325 (~72%) protein-coding genes that were functionally annotated. This genome was found to consist of over 58% repeat sequences, primarily long terminal repeats (LTR-)-retrotransposons. We find an expansion of terpenoid metabolism genes in P. cineraria and its relative Prosopis alba, but not in other legumes. We also observed an amplification of NBS-LRR disease-resistance genes correlated with LTR-associated retrotransposition, and identified 410 retrogenes with an active burst of chimeric retrogene creation that approximately occurred at the same time of divergence of P. cineraria from a common lineage with P. alba~23 Mya. These retrogenes include many biotic defense responses and abiotic stress stimulus responses, as well as the early Nodulin 93 gene. Nodulin 93 gene amplification is consistent with an adaptive response of the species to the low nitrogen in arid desert soil. Consistent with these results, our differentially expressed genes show a tissue specific expression of isoprenoid pathways in shoots, but not in roots, as well as important genes involved in abiotic salt stress in both tissues. Overall, the genome sequence of P. cineraria enriches our understanding of the genomic mechanisms of its disease resistance and abiotic stress tolerance. Thus, it is a very important step in crop and legume improvement.

Genome sequence and genetic diversity analysis of an under-domesticated orphan crop, white fonio (Digitaria exilis).

BACKGROUND: Digitaria exilis, white fonio, is a minor but vital crop of West Africa that is valued for its resilience in hot, dry, and low-fertility environments and for the exceptional quality of its grain for human nutrition. Its success is hindered, however, by a low degree of plant breeding and improvement. FINDINGS: We sequenced the fonio genome with long-read SMRT-cell technology, yielding a ∼761 Mb assembly in 3,329 contigs (N50, 1.73 Mb; L50, 126). The assembly approaches a high level of completion, with a BUSCO score of >99%. The fonio genome was found to be a tetraploid, with most of the genome retained as homoeologous duplications that differ overall by ∼4.3%, neglecting indels. The 2 genomes within fonio were found to have begun their independent divergence ∼3.1 million years ago. The repeat content (>49%) is fairly standard for a grass genome of this size, but the ratio of Gypsy to Copia long terminal repeat retrotransposons (∼6.7) was found to be exceptionally high. Several genes related to future improvement of the crop were identified including shattering, plant height, and grain size. Analysis of fonio population genetics, primarily in Mali, indicated that the crop has extensive genetic diversity that is largely partitioned across a north-south gradient coinciding with the Sahel and Sudan grassland domains. CONCLUSIONS: We provide a high-quality assembly, annotation, and diversity analysis for a vital African crop. The availability of this information should empower future research into further domestication and improvement of fonio.

Integration of high-density genetic mapping with transcriptome analysis uncovers numerous agronomic QTL and reveals candidate genes for the control of tillering in sorghum.

Phenotypes such as branching, photoperiod sensitivity, and height were modified during plant domestication and crop improvement. Here, we perform quantitative trait locus (QTL) mapping of these and other agronomic traits in a recombinant inbred line (RIL) population derived from an interspecific cross between Sorghum propinquum and Sorghum bicolor inbred Tx7000. Using low-coverage Illumina sequencing and a bin-mapping approach, we generated ∼1920 bin markers spanning ∼875 cM. Phenotyping data were collected and analyzed from two field locations and one greenhouse experiment for six agronomic traits, thereby identifying a total of 30 QTL. Many of these QTL were penetrant across environments and co-mapped with major QTL identified in other studies. Other QTL uncovered new genomic regions associated with these traits, and some of these were environment-specific in their action. To further dissect the genetic underpinnings of tillering, we complemented QTL analysis with transcriptomics, identifying 6189 genes that were differentially expressed during tiller bud elongation. We identified genes such as Dormancy Associated Protein 1 (DRM1) in addition to various transcription factors that are differentially expressed in comparisons of dormant to elongating tiller buds and lie within tillering QTL, suggesting that these genes are key regulators of tiller elongation in sorghum. Our study demonstrates the usefulness of this RIL population in detecting domestication and improvement-associated genes in sorghum, thus providing a valuable resource for genetic investigation and improvement to the sorghum community.

Exceptional subgenome stability and functional divergence in the allotetraploid Ethiopian cereal teff.

Teff (Eragrostis tef) is a cornerstone of food security in the Horn of Africa, where it is prized for stress resilience, grain nutrition, and market value. Here, we report a chromosome-scale assembly of allotetraploid teff (variety Dabbi) and patterns of subgenome dynamics. The teff genome contains two complete sets of homoeologous chromosomes, with most genes maintaining as syntenic gene pairs. TE analysis allows us to estimate that the teff polyploidy event occurred ~1.1 million years ago (mya) and that the two subgenomes diverged ~5.0 mya. Despite this divergence, we detect no large-scale structural rearrangements, homoeologous exchanges, or biased gene loss, in contrast to many other allopolyploids. The two teff subgenomes have partitioned their ancestral functions based on divergent expression across a diverse expression atlas. Together, these genomic resources will be useful for accelerating breeding of this underutilized grain crop and for fundamental insights into polyploid genome evolution.

Genome Sequence Analysis of Enterobacter sp. C6, Found in the Pitcher Fluids of Sarracenia rosea.

An unclassified Enterobacter strain was isolated from the pitcher fluids of a Sarracenia rosea pitcher plant growing at Splinter Hill Bog in Alabama. Its genome was sequenced using the Illumina platform. A genome assembly of 4,673,815 bp was obtained. In total, 4,646 protein-encoding sequences and 72 RNA genes are predicted from this assembly.

Draft Genome Sequence of a Serratia marcescens Strain Isolated from the Pitcher Fluids of a Sarracenia Pitcher Plant.

The genome of a Serratia marcescens strain (C7) that was found in the pitcher fluids of a Sarracenia rosea pitcher plant was sequenced using the Illumina platform. A 5,543,750-bp genome assembly was obtained. A total of 6,278 coding sequences are predicted from this assembly.

Phoenix phylogeny, and analysis of genetic variation in a diverse collection of date palm (Phoenix dactylifera) and related species.

Date palm (Phoenix dactylifera), one of the most ancient crops, is grown commercially in >30 countries. Using whole plastome assemblies, phylogenetic analyses revealed that cultivated date palm accessions share the same clade with P hoenix sylvestris, P hoenix pusilla and P hoenix acaulis, which are native to the Indian subcontinent, and Phoenix caespitosa that is native to the Arabian Peninsula and the deserts of Somalia. Analysis of genetic diversity and genetic relationships among date palm accessions from 13 producing countries involved 195 date palm accessions that were genotyped at 19 microsatellite loci. Extensive genetic diversity was observed, with many accessions heterozygous for most markers in this clonally propagated crop. The average number of alleles per locus (42.1), expected heterozygosity (0.8), observed heterozygosity (0.47) and fixation indices (FST = 0.42) demonstrated substantial genetic diversity and population structure. Iraqi accessions were found to have the richest allelic diversity, and the most private alleles. The model-based Bayesian method indicated that these accessions could be broadly divided into two structure groups, one group with predominantly African accessions and another predominantly Asian. Some germplasm, especially from Tunisia and Iraq, deviated from this generalization. Many accessions in the STRUCTURE-derived groups were found to be genetic admixtures, with gene flow between Asian and African groups. Indian and Pakistani date palms were found to be most closely related to North African germplasm.

Species-Associated Differences in the Below-Ground Microbiomes of Wild and Domesticated Setaria.

The rhizosphere microbiome is known to play a crucial role in promoting plant growth, partly by countering soil-borne phytoparasites and by improving nutrient uptake. The abundance and composition of the rhizosphere and root-associated microbiota are influenced by several factors, including plant species and genotype. We hypothesize that crop domestication might influence the composition and diversity of plant-associated microbiomes. We tested the contribution of domestication to the bacterial and archaeal root and soil composition associated with six genotypes of domesticated Setaria italica and four genotypes of its wild ancestor, S. viridis. The bacterial microbiome in the rhizoplane and root endophyte compartments, and the archaea in the endophyte compartment, showed major composition differences. For instance, members of the Betaproteobacteria and Firmicutes were overrepresented in S. italica root samples compared to S. viridis. Metagenomic analysis of samples that contained both root surface-bound (rhizoplane) and inside-root (endophytic) bacteria defined two unique microbial communities only associated with S. italica roots and one only associated with S. viridis roots. Root endophytic bacteria were found in six discernible communities, of which four were primarily on S. italica and two primarily on S. viridis. Among archaea, Methanobacteria, and Methanomicrobia exhibited species-associated differences in the rhizosphere and root compartments, but most detected archaea were not classified more specifically than at the level of phylum. These results indicate a host genetic contribution to the microbial composition in Setaria, and suggest that domestication has selected for specific associations in the root and in the rhizosphere.

Analysis of retrotransposon abundance, diversity and distribution in holocentric Eleocharis (Cyperaceae) genomes.

Background and Aims: Long terminal repeat-retrotransposons (LTR-RTs) comprise a large portion of plant genomes, with massive repeat blocks distributed across the chromosomes. Eleocharis species have holocentric chromosomes, and show a positive correlation between chromosome numbers and the amount of nuclear DNA. To evaluate the role of LTR-RTs in karyotype diversity in members of Eleocharis (subgenus Eleocharis), the occurrence and location of different members of the Copia and Gypsy superfamilies were compared, covering interspecific variations in ploidy levels (considering chromosome numbers), DNA C-values and chromosomal arrangements. Methods: The DNA C-value was estimated by flow cytometry. Genomes of Eleocharis elegans and E. geniculata were partially sequenced using Illumina MiSeq assemblies, which were a source for searching for conserved proteins of LTR-RTs. POL domains were used for recognition, comparing families and for probe production, considering different families of Copia and Gypsy superfamilies. Probes were obtained by PCR and used in fluorescence in situ hybridization (FISH) against chromosomes of seven Eleocharis species. Key Results: A positive correlation between ploidy levels and the amount of nuclear DNA was observed, but with significant variations between samples with the same ploidy levels, associated with repetitive DNA fractions. LTR-RTs were abundant in E. elegans and E. geniculata genomes, with a predominance of Copia Sirevirus and Gypsy Athila/Tat clades. FISH using LTR-RT probes exhibited scattered and clustered signals, but with differences in the chromosomal locations of Copia and Gypsy. The diversity in LTR-RT locations suggests that there is no typical chromosomal distribution pattern for retrotransposons in holocentric chromosomes, except the CRM family with signals distributed along chromatids. Conclusions: These data indicate independent fates for each LTR-RT family, including accumulation between and within chromosomes and genomes. Differential activity and small changes in LTR-RTs suggest a secondary role in nuclear DNA variation, when compared with ploidy changes.

Gene loss and genome rearrangement in the plastids of five Hemiparasites in the family Orobanchaceae.

BACKGROUND: The chloroplast genomes (plastome) of most plants are highly conserved in structure, gene content, and gene order. Parasitic plants, including those that are fully photosynthetic, often contain plastome rearrangements. These most notably include gene deletions that result in a smaller plastome size. The nature of gene loss and genome structural rearrangement has been investigated in several parasitic plants, but their timing and contributions to the adaptation of these parasites requires further investigation, especially among the under-studied hemi-parasites. RESULTS: De novo sequencing, assembly and annotation of the chloroplast genomes of five photosynthetic parasites from the family Orobanchaceae were employed to investigate plastome dynamics. Four had major structural rearrangements, including gene duplications and gene losses, that differentiated the taxa. The facultative parasite Aureolaria virginica had the most similar genome content to its close non-parasitic relative, Lindenbergia philippensis, with similar genome size and organization, and no differences in gene content. In contrast, the facultative parasite Buchnera americana and three obligate parasites in the genus Striga all had enlargements of their plastomes, primarily caused by expansion within the large inverted repeats (IRs) that are a standard plastome feature. Some of these IR increases were shared by multiple investigated species, but others were unique to particular lineages. Gene deletions and pseudogenization were also both shared and lineage-specific, with particularly frequent and independent loss of the ndh genes involved in electron recycling. CONCLUSIONS: Five new plastid genomes were fully assembled and compared. The results indicate that plastome instability is common in parasitic plants, even those that retain the need to perform essential plastid functions like photosynthesis. Gene losses were slow and not identical across taxa, suggesting that different lineages had different uses or needs for some of their plastome gene content, including genes involved in some aspects of photosynthesis. Recent repeat region extensions, some unique to terminal species branches, were observed after the divergence of the Buchnera/Striga clade, suggesting that this otherwise rare event has some special value in this lineage.

Developmental programmed cell death during asymmetric microsporogenesis in holocentric species of Rhynchospora (Cyperaceae).

Members of the Cyperaceae family exhibit an asymmetric microsporogenesis that results in the degeneration of three out of four meiotic products. Efforts have been made previously to describe the resulting structure, named the pseudomonad, but mechanisms concerning the establishment of cell domains, nuclear development, and programmed cell death are largely unknown. Using the Rhynchospora genus as a model, evidence for cell asymmetry, cytoplasmic isolation, and programmed cell death was obtained by a combination of electron microscopic, cytochemical, immunocytochemical, in situ hybridization, and flow cytometric methods. Degenerative cells were identified at the abaxial region, with the cytoskeleton marking their delimitation from the functional domain after meiosis. After attempting to initiate cell division with an unreplicated genome and abnormal spindle assembly, these cells exhibited a gradual process of cytoplasmic contraction associated with hypermethylation of cytosines and differential loss of DNA. These results indicate that the asymmetric tetrad establishes a functional cell, where one nucleus is preferentially selected to survive. Degenerative haploid cells are then eliminated in a multistep process associated with mitotic disorder, non-random elimination of repetitive DNA, vacuolar cell death, and DNA fragmentation.

Salicylic acid treatment and expression of an RNA-dependent RNA polymerase 1 transgene inhibit lethal symptoms and meristem invasion during tobacco mosaic virus infection in Nicotiana benthamiana.

BACKGROUND: Host RNA-dependent RNA polymerases (RDRs) 1 and 6 contribute to antiviral RNA silencing in plants. RDR6 is constitutively expressed and was previously shown to limit invasion of Nicotiana benthamiana meristem tissue by potato virus X and thereby inhibit disease development. RDR1 is inducible by salicylic acid (SA) and several other phytohormones. But although it contributes to basal resistance to tobacco mosaic virus (TMV) it is dispensable for SA-induced resistance in inoculated leaves. The laboratory accession of N. benthamiana is a natural rdr1 mutant and highly susceptible to TMV. However, TMV-induced symptoms are ameliorated in transgenic plants expressing Medicago truncatula RDR1. RESULTS: In MtRDR1-transgenic N. benthamiana plants the spread of TMV expressing the green fluorescent protein (TMV.GFP) into upper, non-inoculated, leaves was not inhibited. However, in these plants exclusion of TMV.GFP from the apical meristem and adjacent stem tissue was greater than in control plants and this exclusion effect was enhanced by SA. TMV normally kills N. benthamiana plants but although MtRDR1-transgenic plants initially displayed virus-induced necrosis they subsequently recovered. Recovery from disease was markedly enhanced by SA treatment in MtRDR1-transgenic plants whereas in control plants SA delayed but did not prevent systemic necrosis and death. Following SA treatment of MtRDR1-transgenic plants, extractable RDR enzyme activity was increased and Western blot analysis of RDR extracts revealed a band cross-reacting with an antibody raised against MtRDR1. Expression of MtRDR1 in the transgenic N. benthamiana plants was driven by a constitutive 35S promoter derived from cauliflower mosaic virus, confirmed to be non-responsive to SA. This suggests that the effects of SA on MtRDR1 are exerted at a post-transcriptional level. CONCLUSIONS: MtRDR1 inhibits severe symptom development by limiting spread of virus into the growing tips of infected plants. Thus, RDR1 may act in a similar fashion to RDR6. MtRDR1 and SA acted additively to further promote recovery from disease symptoms in MtRDR1-transgenic plants. Thus it is possible that SA promotes MtRDR1 activity and/or stability through post-transcriptional effects.

Whole plastome sequences from five ginger species facilitate marker development and define limits to barcode methodology.

Plants from the Zingiberaceae family are a key source of spices and herbal medicines. Species identification within this group is critical in the search for known and possibly novel bioactive compounds. To facilitate precise characterization of this group, we have sequenced chloroplast genomes from species representing five major groups within Zingiberaceae. Generally, the structure of these genomes is similar to the basal angiosperm excepting an expansion of 3 kb associated with the inverted repeat A region. Portions of this expansion appear to be shared across the entire Zingiberales order, which includes gingers and bananas. We used whole plastome alignment information to develop DNA barcodes that would maximize the ability to differentiate species within the Zingiberaceae. Our computation pipeline identified regions of high variability that were flanked by highly conserved regions used for primer design. This approach yielded hitherto unexploited regions of variability. These theoretically optimal barcodes were tested on a range of species throughout the family and were found to amplify and differentiate genera and, in some cases, species. Still, though these barcodes were specifically optimized for the Zingiberaceae, our data support the emerging consensus that whole plastome sequences are needed for robust species identification and phylogenetics within this family.

High-throughput discovery of mutations in tef semi-dwarfing genes by next-generation sequencing analysis.

Tef (Eragrostis tef) is a major cereal crop in Ethiopia. Lodging is the primary constraint to increasing productivity in this allotetraploid species, accounting for losses of ∼15-45% in yield each year. As a first step toward identifying semi-dwarf varieties that might have improved lodging resistance, an ∼6× fosmid library was constructed and used to identify both homeologues of the dw3 semi-dwarfing gene of Sorghum bicolor. An EMS mutagenized population, consisting of ∼21,210 tef plants, was planted and leaf materials were collected into 23 superpools. Two dwarfing candidate genes, homeologues of dw3 of sorghum and rht1 of wheat, were sequenced directly from each superpool with 454 technology, and 120 candidate mutations were identified. Out of 10 candidates tested, six independent mutations were validated by Sanger sequencing, including two predicted detrimental mutations in both dw3 homeologues with a potential to improve lodging resistance in tef through further breeding. This study demonstrates that high-throughput sequencing can identify potentially valuable mutations in under-studied plant species like tef and has provided mutant lines that can now be combined and tested in breeding programs for improved lodging resistance.

De novo genome sequencing and comparative genomics of date palm (Phoenix dactylifera).

Date palm is one of the most economically important woody crops cultivated in the Middle East and North Africa and is a good candidate for improving agricultural yields in arid environments. Nonetheless, long generation times (5-8 years) and dioecy (separate male and female trees) have complicated its cultivation and genetic analysis. To address these issues, we assembled a draft genome for a Khalas variety female date palm, the first publicly available resource of its type for a member of the order Arecales. The ∼380 Mb sequence, spanning mainly gene-rich regions, includes >25,000 gene models and is predicted to cover ∼90% of genes and ∼60% of the genome. Sequencing of eight other cultivars, including females of the Deglet Noor and Medjool varieties and their backcrossed males, identified >3.5 million polymorphic sites, including >10,000 genic copy number variations. A small subset of these polymorphisms can distinguish multiple varieties. We identified a region of the genome linked to gender and found evidence that date palm employs an XY system of gender inheritance.

Characterization of a Brome mosaic virus strain and its use as a vector for gene silencing in monocotyledonous hosts.

Virus-induced gene silencing (VIGS) is used to analyze gene function in dicotyledonous plants but less so in monocotyledonous plants (particularly rice and corn), partially due to the limited number of virus expression vectors available. Here, we report the cloning and modification for VIGS of a virus from Festuca arundinacea Schreb. (tall fescue) that caused systemic mosaic symptoms on barley, rice, and a specific cultivar of maize (Va35) under greenhouse conditions. Through sequencing, the virus was determined to be a strain of Brome mosaic virus (BMV). The virus was named F-BMV (F for Festuca), and genetic determinants that controlled the systemic infection of rice were mapped to RNAs 1 and 2 of the tripartite genome. cDNA from RNA 3 of the Russian strain of BMV (R-BMV) was modified to accept inserts from foreign genes. Coinoculation of RNAs 1 and 2 from F-BMV and RNA 3 from R-BMV expressing a portion of a plant gene to leaves of barley, rice, and maize plants resulted in visual silencing-like phenotypes. The visual phenotypes were correlated with decreased target host transcript levels in the corresponding leaves. The VIGS visual phenotype varied from maintained during silencing of actin 1 transcript expression to transient with incomplete penetration through affected tissue during silencing of phytoene desaturase expression. F-BMV RNA 3 was modified to allow greater accumulation of virus while minimizing virus pathogenicity. The modified vector C-BMV(A/G) (C for chimeric) was shown to be useful for VIGS. These BMV vectors will be useful for analysis of gene function in rice and maize for which no VIGS system is reported.