A universal design of restructured dimer antigens: Development of a superior vaccine against the paramyxovirus in transgenic rice.

The development of vaccines, which induce effective immune responses while ensuring safety and affordability, remains a substantial challenge. In this study, we proposed a vaccine model of a restructured "head-to-tail" dimer to efficiently stimulate B cell response. We also demonstrate the feasibility of using this model to develop a paramyxovirus vaccine through a low-cost rice endosperm expression system. Crystal structure and small-angle X-ray scattering data showed that the restructured hemagglutinin-neuraminidase (HN) formed tetramers with fully exposed quadruple receptor binding domains and neutralizing epitopes. In comparison with the original HN antigen and three traditional commercial whole virus vaccines, the restructured HN facilitated critical epitope exposure and initiated a faster and more potent immune response. Two-dose immunization with 0.5 µg of the restructured antigen (equivalent to one-127th of a rice grain) and one-dose with 5 µg completely protected chickens against a lethal challenge of the virus. These results demonstrate that the restructured HN from transgenic rice seeds is safe, effective, low-dose useful, and inexpensive. We provide a plant platform and a simple restructured model for highly effective vaccine development.

Rice-produced classical swine fever virus glycoprotein E2 with herringbone-dimer design to enhance immune responses.

Pestiviruses, including classical swine fever virus, remain a concern for global animal health and are responsible for major economic losses of livestock worldwide. Despite high levels of vaccination, currently available commercial vaccines are limited by safety concerns, moderate efficacy, and required high doses. The development of new vaccines is therefore essential. Vaccine efforts should focus on optimizing antigen presentation to enhance immune responses. Here, we describe a simple herringbone-dimer strategy for efficient vaccine design, using the classical swine fever virus E2 expressed in a rice endosperm as an example. The expression of rE2 protein was identified, with the rE2 antigen accumulating to 480 mg/kg. Immunological assays in mice, rabbits, and pigs showed high antigenicity of rE2. Two immunizations with 284 ng of the rE2 vaccine or one shot with 5.12 µg provided effective protection in pigs without interference from pre-existing antibodies. Crystal structure and small-angle X-ray scattering results confirmed the stable herringbone dimeric conformation, which had two fully exposed duplex receptor binding domains. Our results demonstrated that rice endosperm is a promising platform for precise vaccine design, and this strategy can be universally applied to other Flaviviridae virus vaccines.

Knock-in at GluA1 locus improves recombinant human serum albumin expression in rice grain.

Improving recombinant protein expression is a perpetual goal for molecular pharming. However, over-transcription of recombinant protein induces ER stress, and causes protein degradation. Here, we describe a knock-in approach to integrate a human serum albumin expression cassette into the locus of the rice storage protein GluA1 by site-specific integration via the nonhomologous end joining (NHEJ) pathway. The expression level of OsrHSA in the knock-in (KI) lines was much higher than that of the random integration (RI) lines. ER stress in KI line endosperm cells was not significantly altered even after massive OsrHSA accumulation in rice endosperm cell. Instead, ER stress induced by high OsrHSA expression was alleviated in the KI line via the inositol-requiring enzyme 1 (IRE1)-mediated/OsbZIP50 pathway. Furthermore, improvement of OsrHSA expression in KI lines is likely due to reduction of contents of glutelin and globulin in rice endosperm cell. These results provide insight into an approach to improving recombinant protein accumulation by alleviating ER stress and protein trafficking.

Studies on the cold tolerance of ratoon 'Chaling' common wild rice.

BACKGROUND: Rice is the staple food of many people around the world. However, most rice varieties, especially widely grown indica varieties and hybrids, are sensitive to cold stress. In order to provide a basis for the utilization of a common wild rice (CWR, Oryza rufipogon Griff.) named 'Chaling' CWR in cold-tolerant rice breeding and deepen the understanding of rice cold tolerance, the cold tolerance of ratoon 'Chaling' CWR was studied under the stress of the natural low temperature in winter in Changsha, Hunan province, China, especially under the stress of abnormal natural low temperature in Changsha in 2008, taking other ratoon CWR accessions and ratoon cultivated rice phenotypes as control. RESULTS: The results showed that ratoon 'Chaling' CWR can safely overwinter under the natural conditions in Changsha (28° 22' N), Hunan province, China, which is a further and colder northern place than its habitat, even if it suffers a long-term low temperature stress with ice and snow. In 2008, an extremely cold winter appeared in Changsha, i.e., the average daily mean temperature of 22 consecutive days from January 13 to February 3 was - 1.0 °C, and the extreme low temperature was - 4.7 °C. After subjected to this long-term cold stress, the overwinter survival rate of ratoon 'Chaling' CWR was 100%, equals to that of ratoon 'Dongxiang' CWR which is northernmost distribution in the word among wild rice populations, higher than those of ratoon 'Fusui' CWR, ratoon 'Jiangyong' CWR, and ratoon 'Liujiang' CWR (63.55-83.5%) as well as those of ratoon 'Hainan' CWR, ratoon 'Hepu' CWR, and all the ratoon cultivated rice phenotypes including 3 japonica ones, 3 javanica ones, and 5 indica ones (0.0%). CONCLUSIONS: The results indicate that ratoon 'Chaling' CWR possesses strong cold tolerance and certain freezing tolerance.

Studies on the cold tolerance of ratoon 'Chaling' common wild rice

BACKGROUND: Rice is the staple food of many people around the world. However, most rice varieties, especially widely grown indica varieties and hybrids, are sensitive to cold stress. In order to provide a basis for the utilization of a common wild rice (CWR, Oryza rufipogon Griff.) named 'Chaling' CWR in cold-tolerant rice breeding and deepen the understanding of rice cold tolerance, the cold tolerance of ratoon 'Chaling' CWR was studied under the stress of the natural low temperature in winter in Changsha, Hunan province, China, especially under the stress of abnormal natural low temperature in Changsha in 2008, taking other ratoon CWR accessions and ratoon cultivated rice phenotypes as control. RESULTS: The results showed that ratoon 'Chaling' CWR can safely overwinter under the natural conditions in Changsha (28° 22' N), Hunan province, China, which is a further and colder northern place than its habitat, even if it suffers a long-term low temperature stress with ice and snow. In 2008, an extremely cold winter appeared in Changsha, i.e., the average daily mean temperature of 22 consecutive days from January 13 to February 3 was - 1.0 °C, and the extreme low temperature was - 4.7 °C. After subjected to this long-term cold stress, the overwinter survival rate of ratoon 'Chaling' CWR was 100%, equals to that of ratoon 'Dongxiang' CWR which is northernmost distribution in the word among wild rice populations, higher than those of ratoon 'Fusui' CWR, ratoon 'Jiangyong' CWR, and ratoon 'Liujiang' CWR (63.55-83.5%) as well as those of ratoon 'Hainan' CWR, ratoon 'Hepu' CWR, and all the ratoon cultivated rice phenotypes including 3 japonica ones, 3 javanica ones, and 5 indica ones (0.0%). CONCLUSIONS: The results indicate that ratoon 'Chaling' CWR possesses strong cold tolerance and certain freezing tolerance.

A 90-day subchronic toxicology screen of genetically modified rice Lac-3 and its effects on the gut microbiota in Sprague-Dawley rats.

A 90-day subchronic toxicology screen of genetically modified (GM) rice Lac-3 expressing human lactoferrin (hLF) and its effects on the gut microbiota were studied in comparison to non-GM rice fed to Sprague-Dawley (SD) rats. Three different dietary concentrations (17.5%, 35% and 70%, w/w) of the GM rice or its corresponding non-GM rice were used. Additionally, the phylotypes of gut microbiota in the control group, the 70% GM rice diet group and the 70% non-GM rice diet group on day 90 were determined by 16S rRNA sequencing. The results of the 90-day subchronic feeding study demonstrated that the GM rice Lac-3 containing human lactoferrin (LF) gene is considered as safe as the non-GM rice. The results of bacterial 16S rRNA sequencing showed that the structure of gut microbiota in the 70% GM group slightly changed when compared with the control group and the 70% non-GM group. There were no significant differences in the microbiota diversity among the three groups.

Assessment of the immunogenicity of residual host cell protein impurities of OsrHSA.

Human serum albumin (HSA) is the most abundant protein in human plasma and is widely used at high doses for treating various diseases. Recombinant HSA is an alternative approach to plasma-derived HSA, providing increased safety and an unlimited supply. However, the safety of the residual host cell proteins (HCPs) co-purified with Oryza sativa HSA (OsrHSA) remains to be determined. An animal system was used to assess the immunogenicity of OsrHSA and its residual HCPs. Low immunogenicity and immunotoxicity of the residual HCPs at a dose of 25 µg/kg, equivalent to 25 times the clinical dosage of HSA, were observed. An anti-drug-antibody (ADA) analysis revealed that anti-HSA, anti-OsrHSA or anti-HCP antibodies developed with a low frequency in pHSA and OsrHSA treatments, but the titers were as low as 1.0-2.0. Furthermore, the titer and the incidence of the specific antibodies were not significantly different between the pHSA and OsrHSA groups, indicating that OsrHSA presents similar immunogenicity to that of pHSA. More importantly, no cytokines were stimulated after the administration of OsrHSA and the residual HCPs, suggesting that there was no risk of a cytokine storm. These results demonstrated that the residual HCPs from OsrHSA have low immunogenicity, indicating that the rice endosperm is one of the best hosts for plant molecular pharming.

OsPRR37 confers an expanded regulation of the diurnal rhythms of the transcriptome and photoperiodic flowering pathways in rice.

The circadian clock enables organisms to rapidly adapt to the ever-changing environmental conditions that are caused by daily light/dark cycles. Circadian clock genes universally affect key agricultural traits, particularly flowering time. Here, we show that OsPRR37, a circadian clock gene, delays rice flowering time in an expression level-dependent manner. Using high-throughput mRNA sequencing on an OsPRR37 overexpressing transgenic line (OsPRR37-OE5) and the recipient parent Guangluai4 that contains the loss-of-function Osprr37, we identify 14,992 genes that display diurnal rhythms, which account for 52.9% of the transcriptome. Overexpressing OsPRR37 weakens the transcriptomic rhythms and alters the phases of rhythmic genes. In total, 3,210 differentially expressed genes (DEGs) are identified, among which 1,863 rhythmic DEGs show a correlation between the change of absolute amplitudes and the mean expression levels. We further reveal that OsPRR37 functions as a transcriptional repressor to repress the expression levels and amplitudes of day-phased clock genes. More importantly, OsPRR37 confers expanded regulation on the evening-phased rhythmic DEGs by repressing the morning-phased rhythmic DEGs. Further study shows that OsPRR37 expands its regulation on flowering pathways by repressing Ehd1. Thus, our results demonstrate an expanded regulation mechanism of the circadian clock on the diurnal rhythms of the transcriptome.

Expression of α-1,6-fucosyltransferase (FUT8) in rice grain and immunogenicity evaluation of plant-specific glycans.

Rice seed is a cost-effective bioreactor for the large-scale production of pharmaceuticals. However, convincing evidence of the immunogenicity of plant-specific glycans is still limited although plant-specific glycans are considered potential allergic antigens. In the present study, we found that the α-1,3-fucose content of the glycoprotein produced from rice seed was much lower than that in leaf, and conversely, a higher ß-1,2-xylose content was detected in seed than that in leaf. We detected the α-1,6-fucose content in the glutelin and recombinant human α1-antitrypsin (OsrAAT). The further results in a line containing AAT and FUT8 genes indicated that the α-1,6-fucose content of modified glycosylated recombinant α1-antitrypsin (mgOsrAAT) was 38.4%, while glutelin was only 6.8%. Interestingly, the α-1,3-fucose content of mgOsrAAT was significantly reduced by 59.8% compared with that of OsrAAT. Furthermore, we assessed the immunogenicity of OsrAAT, mgOsrAAT and human α1-antitrypsin (hAAT) using an animal system. The PCA results indicated no significant differences in the IgG, IgM and IgE titers among OsrAAT, mgOsrAAT and hAAT. Further studies revealed that those antibodies were mainly from α-1,3-fucose, but not from ß-1,2-xylose, indicating that α-1,3-fucose was the major immunogenic resource. Our results demonstrated that α-1,3-fucose contents in seed proteins was much less than that of leaf, and could not be a plant-specific glycan because it also exists in human proteins.

The rice DUF1620-containing and WD40-like repeat protein is required for the assembly of the restoration of fertility complex.

Cytoplasmic male sterility (CMS) and restoration of fertility (Rf) are widely distributed in plant species utilized by humans. RF5 and GRP162 are subunits of the restoration of fertility complex (RFC) in Hong-Lian rice. Despite the fact that the RFC is 400-500 kDa in size, the other proteins or factors in the complex still remain unknown. Here, we identified RFC subunit 3, which encodes a DUF1620-containing and WD40-like repeat protein (RFC3) that is present in all tissues but highly expressed in leaves. We established that RFC3 interacts with both RF5 and GRP162 in vitro and in vivo, and is transported into the mitochondria as a membrane protein. Furthermore, CMS RNA (atp6-orfH79) and CMS cytotoxic protein (ORFH79) accumulate when RFC3 is silenced in restorer lines. We presented the analysis with blue-native polyacrylamide gel electrophoresis, indicating that RFC is disrupted in the RNAi line. We concluded that RCF3 is indispensable as a scaffold protein for the assembly of the RFC complex. We unveil a new molecular player of the RFC in the Rf pathway in rice and propose the model of RFC based on these data.

OsPRR37 and Ghd7 are the major genes for general combining ability of DTH, PH and SPP in rice.

Artificial selection of high yield crops and better livestock is paramount importance in breeding programs. Selection of elite parents with preferred traits from a phalanx of inbred lines is extremely laborious, time-consuming and highly random. General combining ability (GCA) was proposed and has been widely used for the evaluation of parents in hybrid breeding for more than half a century. However, the genetic and molecular basis of GCA has been largely overlooked. Here, we present two pleotropic QTLs are accounting for GCA of days to heading (DTH), plant height (PH) and spikelet per panicle (SPP) using an F2-based NCII design, the BC3F2 population as well as a set of nearly isogenic lines (NILs) with five testers. Both GCA1 and GCA2 were loss-of-function gene in low-GCA parent and gain-of-function gene in high-GCA parent, encoding the putative Pseudo-Response Regulators, OsPRR37 and Ghd7, respectively. Overexpression of GCA1 in low-GCA parent significantly increases GCA effects in three traits. Our results demonstrate that two GCA loci associate with OsPRR37 and Ghd7 and reveal that the genes responsible for important agronomic traits could simultaneously account for GCA effects.

Global epigenomic analysis indicates that epialleles contribute to Allele-specific expression via Allele-specific histone modifications in hybrid rice.

BACKGROUND: For heterozygous genes, alleles on the chromatin from two different parents exhibit histone modification variations known as allele-specific histone modifications (ASHMs). The regulation of allele-specific gene expression (ASE) by ASHMs has been reported in animals. However, to date, the regulation of ASE by ASHM genes remains poorly understood in higher plants. RESULTS: We used chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) to investigate the global ASHM profiles of trimethylation on histone H3 lysine 27 (H3K27me3) and histone H3 lysine 36 (H3K36me3) in two rice F1 hybrids. A total of 522 to 550 allele-specific H3K27me3 genes and 428 to 494 allele-specific H3K36me3 genes were detected in GL × 93-11 and GL × TQ, accounting for 11.09% and 26.13% of the total analyzed genes, respectively. The epialleles between parents were highly related to ASHMs. Further analysis indicated that 52.48% to 70.40% of the epialleles were faithfully inherited by the F1 hybrid and contributed to 33.18% to 46.55% of the ASHM genes. Importantly, 66.67% to 82.69% of monoallelic expression genes contained the H3K36me3 modification. Further studies demonstrated a significant positive correlation of ASE with allele-specific H3K36me3 but not with H3K27me3, indicating that ASHM-H3K36me3 primarily regulates ASE in this study. CONCLUSIONS: Our results demonstrate that epialleles from parents can be inherited by the F1 to produce ASHMs in the F1 hybrid. Our findings indicate that ASHM-H3K36me3, rather than H3K27me3, mainly regulates ASE in hybrid rice.

The OsSec18 complex interacts with P0(P1-P2)2 to regulate vacuolar morphology in rice endosperm cell.

BACKGROUND: Sec18p/N-ethylmaleimide-sensitive factor (NSF) is a conserved eukaryotic ATPase, which primarily functions in vesicle membrane fusion from yeast to human. However, the function of the OsSec18 gene, a homologue of NSF in rice, remains unknown. RESULTS: In the present study, we investigated the function of OsSec18 in rice and found that OsSec18 complements the temperature-sensitive phenotype and interferes with vacuolar morphogenesis in yeast. Overexpression of OsSec18 in rice decreased the plant height and 1000-grain weight and altered the morphology of the protein bodies. Further examination revealed that OsSec18 presented as a 290-kDa complex in rice endosperm cells. Moreover, Os60sP0 was identified a component of this complex, demonstrating that the OsSec18 complex contains another complex of P0(P1-P2)2 in rice endosperm cells. Furthermore, we determined that the N-terminus of OsSec18 can interact with the N- and C-termini of Os60sP0, whereas the C-terminus of OsSec18 can only interact with the C-terminus of Os60sP0. CONCLUSION: Our results revealed that the OsSec18 regulates vacuolar morphology in both yeast and rice endosperm cell and the OsSec18 interacts with P0(P1-P2)2 complex in rice endosperm cell.

A SNP in OsMCA1 responding for a plant architecture defect by deactivation of bioactive GA in rice.

Plant architecture directly affects biomass in higher plants, especially grain yields in agricultural crops. In this study, we characterized a recessive mutant, plant architecture determinant (pad), derived from the Oryza sativa ssp. indica cultivar MH86. The mutant exhibited severe dwarf phenotypes, including shorter and stunted leaves, fewer secondary branches during both the vegetative and reproductive growth stages. Cytological studies revealed that pad mutant growth defects are primarily due to the inhibition of cell expansion. The PAD gene was isolated using a map-based cloning strategy. It encodes a plasma membrane protein OsMCA1 and a SNP responsible for a single amino acid change was found in the mutant. PAD was universally expressed in rice tissues from the vegetative to reproductive growth stages, especially in seedlings, nodes and rachillae. Quantitative real-time PCR analysis revealed that the most of the genes responding to gibberellin (GA) metabolism were up-regulated in pad mutant internodes. The endogenous GA content measurement revealed that the levels of GA1 were significantly decreased in the third internode of pad mutants. Moreover, a GA response assay suggested that OsMCA1/PAD might be involved in the regulation of GA metabolism and signal transduction. Our results revealed the pad is a loss-of-function mutant of the OsMCA1/PAD, leading to upregulation of genes related to GA deactivation, which decreased bioactive GA levels.

Fast-tracking determination of homozygous transgenic lines and transgene stacking using a reliable quantitative real-time PCR assay.

The selection of homozygous lines is a crucial step in the characterization of newly generated transgenic plants. This is particularly time- and labor-consuming when transgenic stacking is required. Here, we report a fast and accurate method based on quantitative real-time PCR with a rice gene RBE4 as a reference gene for selection of homozygous lines when using multiple transgenic stacking in rice. Use of this method allowed can be used to determine the stacking of up to three transgenes within four generations. Selection accuracy reached 100 % for a single locus and 92.3 % for two loci. This method confers distinct advantages over current transgenic research methodologies, as it is more accurate, rapid, and reliable. Therefore, this protocol could be used to efficiently select homozygous plants and to expedite time- and labor-consuming processes normally required for multiple transgene stacking. This protocol was standardized for determination of multiple gene stacking in molecular breeding via marker-assisted selection.

Immunotoxicity assessment of rice-derived recombinant human serum albumin using human peripheral blood mononuclear cells.

Human serum albumin (HSA) is extensively used in clinics to treat a variety of diseases, such as hypoproteinemia, hemorrhagic shock, serious burn injuries, cirrhotic ascites and fetal erythroblastosis. To address supply shortages and high safety risks from limited human donors, we recently developed recombinant technology to produce HSA from rice endosperm. To assess the risk potential of HSA derived from Oryza sativa (OsrHSA) before a First-in-human (FIH) trial, we compared OsrHSA and plasma-derived HSA (pHSA), evaluating the potential for an immune reaction and toxicity using human peripheral blood mononuclear cells (PBMCs). The results indicated that neither OsrHSA nor pHSA stimulated T cell proliferation at 1x and 5x dosages. We also found no significant differences in the profiles of the CD4(+) and CD8(+) T cell subsets between OsrHSA- and pHSA-treated cells. Furthermore, the results showed that there were no significant differences between OsrHSA and pHSA in the production of cytokines such as interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), interleukin (IL)-10 and IL-4. Our results demonstrated that OsrHSA has equivalent immunotoxicity to pHSA when using the PBMC model. Moreover, this ex vivo system could provide an alternative approach to predict potential risks in novel biopharmaceutical development.

The phenotypic predisposition of the parent in F1 hybrid is correlated with transcriptome preference of the positive general combining ability parent.

BACKGROUND: Sprague and Tatum (1942) introduced the concepts of general combining ability (GCA) and specific combining ability (SCA) to evaluate the breeding parents and F1 hybrid performance, respectively. Since then, the GCA was widely used in cross breeding for elite parent selection. However, the molecular basis of GCA remains to unknown. RESULTS: We studied the transcriptomes of three varieties and three F1 hybrids using RNA-Sequencing. Transcriptome sequence analysis revealed that the transcriptome profiles of the F1s were similar to the positive GCA-effect parent. Moreover, the expression levels of most differentially expressed genes (DEGs) were equal to the parent with a positive GCA effect. Analysis of the gene expression patterns of gibberellic acid (GA) and flowering time pathways that determine plant height and flowering time in rice validated the preferential transcriptome expression of the parents with positive GCA effect. Furthermore, H3K36me3 modification bias in the Pseudo-Response Regulators (PRR) gene family was observed in the positive GCA effect parents and demonstrated that the phenotype and transcriptome bias in the positive GCA effect parents have been epigenetically regulated by either global modification or specific signaling pathways in rice. CONCLUSIONS: The results revealed that the transcriptome profiles and DEGs in the F1s were highly related to phenotype bias to the positive GCA-effect parent. The transcriptome bias toward high GCA parents in F1 hybrids attributed to H3K36me3 modification both on global modification level and specific signaling pathways. Our results indicated the transcriptome profile and epigenetic modification level bias to high GCA parents could be the molecular basis of GCA.

Quantitation of the residual DNA from rice-derived recombinant human serum albumin.

Residual DNA in recombinant protein pharmaceuticals can potentially cause safety issues in clinical applications; thus, maximum residual limit has been established by drug safety authorities. Assays for residual DNA in Escherichia coli, yeast, and Chinese hamster ovary (CHO) cell expression systems have been established, but no rice residual DNA assay for rice expression systems has been designed. To develop an assay for the quantification of residual DNA that is produced from rice seed, we established a sensitive assay using quantitative real-time polymerase chain reaction (qPCR) based on the 5S ribosomal RNA (rRNA) genes. We found that a 40-cycle qPCR exhibited a linear response when the template concentration was in the range of 2×10(4) to 0.2pg of DNA per reaction in TaqMan and SYBR Green I assays. The amplification efficiency was 103 to 104%, and the amount of residual DNA from recombinant human serum albumin from Oryza sativa (OsrHSA) was less than 3.8ng per dosage, which was lower than that recommended by the World Health Organization (WHO). Our results indicate that the current purification protocol could efficiently remove residual DNA during manufacturing and processing. Furthermore, this protocol could be viable in other cereal crop endosperm expression systems for developing a residual DNA quantitation assay using the highly conserved 5S rRNA gene of the crops.

Deep transcriptome sequencing of rhizome and aerial-shoot in Sorghum propinquum.

Transcriptomic data for Sorghum propinquum, the wild-type sorghum, are limited in public databases. S. propinquum has a subterranean rhizome and transcriptome data will help in understanding the molecular mechanisms underlying rhizome formation. We sequenced the transcriptome of S. propinquum aerial-shoot and rhizome using an Illumina platform. More than 70 % of the genes in the S. propinquum genome were expressed in aerial-shoot and rhizome. The expression patterns of 1963 and 599 genes, including transcription factors, were specific or enriched in aerial-shoot and rhizome respectively, indicating their possible roles in physiological processes in these tissues. Comparative analysis revealed several cis-elements, ACGT box, GCCAC, GATC and TGACG box, which showed significantly higher abundance in aerial-shoot-specific genes. In rhizome-specific genes MYB and ROOTMOTIFTAPOX1 motifs, and 10 promoter and cytokinin-responsive elements were highly enriched. Of the S. propinquum genes, 27.9 % were identified as alternatively spliced and about 60 % of the alternative splicing (AS) events were tissue-specific, suggesting that AS played a crucial role in determining tissue-specific cellular function. The transcriptome data, especially the co-localized rhizome-enriched expressed transcripts that mapped to the publicly available rhizome-related quantitative trait loci, will contribute to gene discovery in S. propinquum and to functional studies of the sorghum genome. Deep transcriptome sequencing revealed a clear difference in the expression patterns of genes between aerial-shoot and rhizome in S. propinquum. This data set provides essential information for future studies into the molecular genetic mechanisms involved in rhizome formation.

Global RNA sequencing reveals that genotype-dependent allele-specific expression contributes to differential expression in rice F1 hybrids.

BACKGROUND: Extensive studies on heterosis in plants using transcriptome analysis have identified differentially expressed genes (DEGs) in F1 hybrids. However, it is not clear why yield in heterozygotes is superior to that of the homozygous parents or how DEGs are produced. Global allele-specific expression analysis in hybrid rice has the potential to answer these questions. RESULTS: We report a genome-wide allele-specific expression analysis using RNA-sequencing technology of 3,637-3,824 genes from three rice F1 hybrids. Of the expressed genes, 3.7% exhibited an unexpected type of monoallelic expression and 23.8% showed preferential allelic expression that was genotype-dependent in reciprocal crosses. Those genes exhibiting allele-specific expression comprised 42.4% of the genes differentially expressed between F1 hybrids and their parents. Allele-specific expression accounted for 79.8% of the genes displaying more than a 10-fold expression level difference between an F1 and its parents, and almost all (97.3%) of the genes expressed in F1, but non-expressed in one parent. Significant allelic complementary effects were detected in the F1 hybrids of rice. CONCLUSIONS: Analysis of the allelic expression profiles of genes at the critical stage for highest biomass production from the leaves of three different rice F1 hybrids identified genotype-dependent allele-specific expression genes. A cis-regulatory mechanism was identified that contributes to allele-specific expression, leading to differential gene expression and allelic complementary effects in F1 hybrids.