Autoploid origin and rapid diploidization of the tetraploid Thinopyrum elongatum revealed by genome differentiation and chromosome pairing in meiosis.

Polyploidy is a common mode of evolution in flowering plants. Both the natural tetraploid Thinopyrum elongatum and the diploid one from the same population show a diploid-like pairing in meiosis. However, debate on the chromosome composition and origin of the tetraploid Th. elongatum is ongoing. In the present study, we obtained the induced tetraploid Th. elongatum and found that the induced and natural tetraploids are morphologically close, except for slower development and lower seed setting. Using probes developed from single chromosome microdissection and a Fosmid library, obvious differentiations were discovered between two chromosome sets (E1 and E2 ) of the natural tetraploid Th. elongatum but not the induced one. Interestingly, hybrid F1 derived from the two different wheat-tetraploid Th. elongatum amphiploids 8802 and 8803 produced seeds well. More importantly, analysis of meiosis in F2 individuals revealed that chromosomes from E1 and E2 could pair well on the durum wheat background with the presence of Ph1. No chromosome set differentiation on the FISH level was discovered from the S1 to S4 generations in the induced one. In metaphase of the meiosis first division in the natural tetraploid, more pairings were bivalents and fewer quadrivalents with ratio of 13.94 II + 0.03 IV (n = 31). Chromosome pairing configuration in the induced tetraploid is 13.05 II + 0.47 IV (n = 19), with the quadrivalent ratio being only slightly higher than the ratio in the natural tetraploid. Therefore, the natural tetraploid Th. elongatum is of autoploid origin and the induced tetraploid Th. elongatum evolutionarily underwent rapid diploidization in the low generation.

Transcriptomic analysis reveals the regulatory mechanisms of messenger RNA (mRNA) and long non-coding RNA (lncRNA) in response to waterlogging stress in rye (Secale cereale L.).

BACKGROUND: Waterlogging stress (WS) negatively impacts crop growth and productivity, making it important to understand crop resistance processes and discover useful WS resistance genes. In this study, rye cultivars and wild rye species were subjected to 12-day WS treatment, and the cultivar Secale cereale L. Imperil showed higher tolerance. Whole transcriptome sequencing was performed on this cultivar to identify differentially expressed (DE) messenger RNAs (DE-mRNAs) and long non-coding RNAs (DE-lncRNAs) involved in WS response. RESULTS: Among the 6 species, Secale cereale L. Imperil showed higher tolerance than wild rye species against WS. The cultivar effectively mitigated oxidative stress, and regulated hydrogen peroxide and superoxide anion. A total of 728 DE-mRNAs and 60 DE-lncRNAs were discovered. Among these, 318 DE-mRNAs and 32 DE-lncRNAs were upregulated, and 410 DE-mRNAs and 28 DE-lncRNAs were downregulated. GO enrichment analysis discovered metabolic processes, cellular processes, and single-organism processes as enriched biological processes (BP). For cellular components (CC), the enriched terms were membrane, membrane part, cell, and cell part. Enriched molecular functions (MF) terms were catalytic activity, binding, and transporter activity. LncRNA and mRNA regulatory processes were mainly related to MAPK signaling pathway-plant, plant hormone signal transduction, phenylpropanoid biosynthesis, anthocyanin biosynthesis, glutathione metabolism, ubiquitin-mediated proteolysis, ABC transporter, Cytochrome b6/f complex, secondary metabolite biosynthesis, and carotenoid biosynthesis pathways. The signalling of ethylene-related pathways was not mainly dependent on AP2/ERF and WRKY transcription factors (TF), but on other factors. Photosynthetic activity was active, and carotenoid levels increased in rye under WS. Sphingolipids, the cytochrome b6/f complex, and glutamate are involved in rye WS response. Sucrose transportation was not significantly inhibited, and sucrose breakdown occurs in rye under WS. CONCLUSIONS: This study investigated the expression levels and regulatory functions of mRNAs and lncRNAs in 12-day waterlogged rye seedlings. The findings shed light on the genes that play a significant role in rye ability to withstand WS. The findings from this study will serve as a foundation for further investigations into the mRNA and lncRNA WS responses in rye.

Genome evolution during bread wheat formation unveiled by the distribution dynamics of SSR sequences on chromosomes using FISH.

BACKGROUND: During the bread wheat speciation by polyploidization, a series of genome rearrangement and sequence recombination occurred. Simple sequence repeat (SSR) sequences, predominately located in heterochromatic regions of chromosomes, are the effective marker for tracing the genomic DNA sequence variations. However, to date the distribution dynamics of SSRs on chromosomes of bread wheat and its donors, including diploid and tetraploid Triticum urartu, Aegilops speltoides, Aegilops tauschii, Triticum turgidum ssp. dicocoides, reflecting the genome evolution events during bread wheat formation had not been comprehensively investigated. RESULTS: The genome evolution was studied by comprehensively comparing the distribution patterns of (AAC)n, (AAG)n, (AGC)n and (AG)n in bread wheat Triticum aestivum var. Chinese Spring and its progenitors T. urartu, A. speltoides, Ae. tauschii, wild tetroploid emmer wheat T. dicocoides, and cultivated emmer wheat T. dicoccum. Results indicated that there are specific distribution patterns in different chromosomes from different species for each SSRs. They provided efficient visible markers for identification of some individual chromosomes and SSR sequence evolution tracing from the diploid progenitors to hexaploid wheat. During wheat speciation, the SSR sequence expansion occurred predominately in the centromeric and pericentromeric regions of B genome chromosomes accompanied by little expansion and elimination on other chromosomes. This result indicated that the B genome might be more sensitive to the "genome shock" and more changeable during wheat polyplodization. CONCLUSIONS: During the bread wheat evolution, SSRs including (AAC)n, (AAG)n, (AGC)n and (AG)n in B genome displayed the greatest changes (sequence expansion) especially in centromeric and pericentromeric regions during the polyploidization from Ae. speltoides S genome, the most likely donor of B genome. This work would enable a better understanding of the wheat genome formation and evolution and reinforce the viewpoint that B genome was originated from S genome.

De Novo Centromere Formation and Centromeric Sequence Expansion in Wheat and its Wide Hybrids.

Centromeres typically contain tandem repeat sequences, but centromere function does not necessarily depend on these sequences. We identified functional centromeres with significant quantitative changes in the centromeric retrotransposons of wheat (CRW) contents in wheat aneuploids (Triticum aestivum) and the offspring of wheat wide hybrids. The CRW signals were strongly reduced or essentially lost in some wheat ditelosomic lines and in the addition lines from the wide hybrids. The total loss of the CRW sequences but the presence of CENH3 in these lines suggests that the centromeres were formed de novo. In wheat and its wide hybrids, which carry large complex genomes or no sequenced genome, we performed CENH3-ChIP-dot-blot methods alone or in combination with CENH3-ChIP-seq and identified the ectopic genomic sequences present at the new centromeres. In adcdition, the transcription of the identified DNA sequences was remarkably increased at the new centromere, suggesting that the transcription of the corresponding sequences may be associated with de novo centromere formation. Stable alien chromosomes with two and three regions containing CRW sequences induced by centromere breakage were observed in the wheat-Th. elongatum hybrid derivatives, but only one was a functional centromere. In wheat-rye (Secale cereale) hybrids, the rye centromere-specific sequences spread along the chromosome arms and may have caused centromere expansion. Frequent and significant quantitative alterations in the centromere sequence via chromosomal rearrangement have been systematically described in wheat wide hybridizations, which may affect the retention or loss of the alien chromosomes in the hybrids. Thus, the centromere behavior in wide crosses likely has an important impact on the generation of biodiversity, which ultimately has implications for speciation.

Identification and Functional Analysis of the psaD Promoter of Chlorella vulgaris Using Heterologous Model Strains.

Chlorella has great potential as a bio-factory for production of value-added compounds. To produce the desired chemicals more efficiently in Chlorella, genetic tools for modification of Chlorella need to be developed, especially an endogenous promoter. In this study, the promoter of photosystem I protein D (psaD) from Chlorella vulgaris UTEX395 was identified. Computational analysis revealed the presence of several putative cis-acting elements, including a potential core element, and light-responsive or stress-responsive elements. Gene expression analysis in heterologous expression system in Chlamydomonasreinhardtii and Nicotianabenthamiana showed that CvpsaD promoter can be used to drive the expression of genes. Functional analysis of this promoter suggested that the initiator element (Inr) is important for its function (i.e., TATA-less promoter) and that an additional factor (e.g., downstream of the transcriptional start site) might be needed for light response. We have shown that the CvpsaD promoter is functional, but not sufficiently strong, both in microalgae and higher plant.

Newly Identified Essential Amino Acids Affecting Chlorella ellipsoidea DGAT1 Function Revealed by Site-Directed Mutagenesis.

Diacylglycerol acyltransferase (DGAT) is a rate-limiting enzyme in the synthesis of triacylglycerol (TAG), the most important form of energy storage in plants. Some residues have previously been proven to be crucial for DGAT1 activity. In this study, we used site-directed mutagenesis of the CeDGAT1 gene from Chlorella ellipsoidea to alter 16 amino acids to investigate effects on DGAT1 function. Of the 16 residues (L482R, E542R, Y553A, G577R, R579D, Y582R, R596D, H603D, H609D, A624R, F629R, S632A, W650R, A651R, Q658H, and P660R), we newly identified 5 (L482, R579, H603, A651, and P660) as being essential for DGAT1 function and 7 (E542, G577, R596, H609, A624, S632, and Q658) that significantly affect DGAT1 function to different degrees, as revealed by heterologous expression of the mutants in yeast strain INVSc1. Importantly, compared with CeDGAT1, expression of the mutant CeDGAT1Y553A significantly increased the total fatty acid and TAG contents of INVSc1. Comparison among CeDGAT1Y553A, GmDGAT1Y341A, AtDGAT1Y364A, BnDGAT1Y347A, and BoDGAT1Y352A, in which tyrosine at the position corresponding to the 553rd residue in CeDGAT1 is changed into alanine, indicated that the impact of changing Y to A at position 553 is specific for CeDGAT1. Overall, the results provide novel insight into the structure and function of DGAT1, as well as a mutant gene with high potential for lipid improvement in microalgae and plants.

Identification and characterization of an efficient acyl-CoA: diacylglycerol acyltransferase 1 (DGAT1) gene from the microalga Chlorella ellipsoidea.

BACKGROUND: Oil in the form of triacylglycerols (TAGs) is quantitatively the most important storage form of energy for eukaryotic cells. Diacylglycerol acyltransferase (DGAT) is considered the rate-limiting enzyme for TAG accumulation. Chlorella, a unicellular eukaryotic green alga, has attracted much attention as a potential feedstock for renewable energy production. However, the function of DGAT1 in Chlorella has not been reported. RESULTS: A full-length cDNA encoding a putative diacylglycerol acyltransferase 1 (DGAT1, EC 2.3.1.20) was obtained from Chlorella ellipsoidea. The 2,142 bp open reading frame of this cDNA, designated CeDGAT1, encodes a protein of 713 amino acids showing no more than 40% identity with DGAT1s of higher plants. Transcript analysis showed that the expression level of CeDGAT1 markedly increased under nitrogen starvation, which led to significant triacylglycerol (TAG) accumulation. CeDGAT1 activity was confirmed in the yeast quadruple mutant strain H1246 by restoring its ability to produce TAG. Upon expression of CeDGAT1, the total fatty acid content in wild-type yeast (INVSc1) increased by 142%, significantly higher than that transformed with DGAT1s from higher plants, including even the oil crop soybean. The over-expression of CeDGAT1 under the NOS promoter in wild-type Arabidopsis thaliana and Brassica napus var. Westar significantly increased the oil content by 8-37% and 12-18% and the average 1,000-seed weight by 9-15% and 6-29%, respectively, but did not alter the fatty acid composition of the seed oil. The net increase in the 1,000-seed total lipid content was up to 25-50% in both transgenic Arabidopsis and B. napus. CONCLUSIONS: We identified a gene encoding DGAT1 in C. ellipsoidea and confirmed that it plays an important role in TAG accumulation. This is the first functional analysis of DGAT1 in Chlorella. This information is important for understanding lipid synthesis and accumulation in Chlorella and for genetic engineering to enhance oil production in microalgae and oil plants.

Isolation and application of P genome-specific DNA sequences of Agropyron Gaertn. in Triticeae.

MAIN CONCLUSION: Different types of P genome sequences and markers were developed, which could be used to analyze the evolution of P genome in Triticeae and identify precisely wheat- A. cristatum introgression lines. P genome of Agropyron Gaertn. plays an important role in Triticeae and could provide many desirable genes conferring high yield, disease resistance, and stress tolerance for wheat genetic improvement. Therefore, it is significant to develop specific sequences and functional markers of P genome. In this study, 126 sequences were isolated from the degenerate oligonucleotide primed-polymerase chain reaction (DOP-PCR) products of microdissected chromosome 6PS. Forty-eight sequences were identified as P genome-specific sequences by dot-blot hybridization and DNA sequences analysis. Among these sequences, 22 displayed the characteristics of retrotransposons, nine and one displayed the characteristics of DNA transposons and tandem repetitive sequence, respectively. Fourteen of 48 sequences were determined to distribute on different regions of P genome chromosomes by fluorescence in situ hybridization, and the distributing regions were as following: all over P genome chromosomes, centromeres, pericentromeric regions, distal regions, and terminal regions. We compared the P genome sequences with other genome sequences of Triticeae and found that the similar sequences of the P genome sequences were widespread in Triticeae, but differentiation occurred to various extents. Additionally, thirty-four molecular markers were developed from the P genome sequences, which could be used for analyzing the evolutionary relationship among 16 genomes of 18 species in Triticeae and identifying P genome chromatin in wheat-A. cristatum introgression lines. These results will not only facilitate the study of structure and evolution of P genome chromosomes, but also provide a rapid detecting tool for effective utilization of desirable genes of P genome in wheat improvement.

Effects of drought and salt-stresses on gene expression in Caragana korshinskii seedlings revealed by RNA-seq.

BACKGROUND: Drought and soil salinity are major abiotic stresses. The mechanisms of stress tolerance have been studied extensively in model plants. Caragana korshinskii is characterized by high drought and salt tolerance in northwestern China; unique patterns of gene expression allow it to tolerate the stress imposed by dehydration and semi-desert saline soil. There have, however, been no reports on the differences between C. korshinskii and model plants in the mechanisms underlying their drought and salt tolerance and regulation of gene expression. RESULTS: Three sequencing libraries from drought and salt-treated whole-seedling- plants and the control were sequenced to investigate changes in the C. korshinskii transcriptome in response to drought and salt stresses. Of the 129,451 contigs, 70,662 (54.12 %) were annotated with gene descriptions, gene ontology (GO) terms, and metabolic pathways, with a cut-off E-value of 10(-5). These annotations included 56 GO terms, 148 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and 25 Clusters of Orthologous Groups (COG). On comparison of the transcriptomes of the control, drought- and salt-treated plants, 1630 and 1521 contigs showed significant differences in transcript abundance under drought and salt stresses. Compared to the differentially expressed genes (DEGs) in drought- or salt-treated Arabidopsis in the database, 542 DEGs in drought-treated C. korshinskii and 529 DEGs in salt-treated samples were presumably unique to C. korshinskii. The transcription profiles revealed that genes related to transcription factors, protein kinases, and antioxidant enzymes are relevant to the tolerance of drought and salt stress in this species. The expression patterns of 38 randomly selected DEGs were confirmed by quantitative real-time PCR and were essentially consistent with the changes in transcript abundance identified by RNA-seq. CONCLUSIONS: The present study identified potential genes involved in drought and salt tolerance in C. korshinskii, as well as many DEGs uniquely expressed in drought- or salt-treated C. korshinskii samples compared to Arabidopsis. To our knowledge, this study is the first exploration of the C. korshinskii transcriptome under drought and salt conditions, and these results will facilitate the discovery of specific stress-resistance-related genes in C. korshinskii, possibly leading to the development of novel plant cultivars through genetic engineering.

Genomic Foundation of Starch-to-Lipid Switch in Oleaginous Chlorella spp.

The ability to rapidly switch the intracellular energy storage form from starch to lipids is an advantageous trait for microalgae feedstock. To probe this mechanism, we sequenced the 56.8-Mbp genome of Chlorella pyrenoidosa FACHB-9, an industrial production strain for protein, starch, and lipids. The genome exhibits positive selection and gene family expansion in lipid and carbohydrate metabolism and genes related to cell cycle and stress response. Moreover, 10 lipid metabolism genes might be originated from bacteria via horizontal gene transfer. Transcriptomic dynamics tracked via messenger RNA sequencing over six time points during metabolic switch from starch-rich heterotrophy to lipid-rich photoautotrophy revealed that under heterotrophy, genes most strongly expressed were from the tricarboxylic acid cycle, respiratory chain, oxidative phosphorylation, gluconeogenesis, glyoxylate cycle, and amino acid metabolisms, whereas those most down-regulated were from fatty acid and oxidative pentose phosphate metabolism. The shift from heterotrophy into photoautotrophy highlights up-regulation of genes from carbon fixation, photosynthesis, fatty acid biosynthesis, the oxidative pentose phosphate pathway, and starch catabolism, which resulted in a marked redirection of metabolism, where the primary carbon source of glycine is no longer supplied to cell building blocks by the tricarboxylic acid cycle and gluconeogenesis, whereas carbon skeletons from photosynthesis and starch degradation may be directly channeled into fatty acid and protein biosynthesis. By establishing the first genetic transformation in industrial oleaginous C. pyrenoidosa, we further showed that overexpression of an NAD(H) kinase from Arabidopsis (Arabidopsis thaliana) increased cellular lipid content by 110.4%, yet without reducing growth rate. These findings provide a foundation for exploiting the metabolic switch in microalgae for improved photosynthetic production of food and fuels.

Small GTP-binding protein PdRanBP regulates vascular tissue development in poplar.

BACKGROUND: Previous research has demonstrated that ectopic expression of Ran-binding protein (RanBP) in Arabidopsis results in more axillary buds and reduced apical dominance compared to WT plants. However, the function of RanBP in poplar, which has very typical secondary growth, remains unclear. Here, the Populus deltoides (Marsh.) RanBP gene (PdRanBP) was isolated and functionally characterized by ectopic expression in a hybrid poplar (P. davidiana Dode × P. bolleana Lauche). RESULTS: PdRanBP was predominantly expressed in leaf buds and tissues undergoing secondary wall expansion, including immature xylem and immature phloem in the stem. Overexpression of PdRanBP in poplar increased the number of sylleptic branches and the proportion of cells in the G2 phase of the cell cycle, retarded plant growth, consistently decreased the size of the secondary xylem and secondary phloem zones, and reduced the expression levels of cell wall biosynthesis genes. The downregulation of PdRanBP facilitated secondary wall expansion and increased stem height, the sizes of the xylem and phloem zones, and the expression levels of cell wall biosynthesis genes. CONCLUSIONS: These results suggest that PdRanBP influences the apical and radial growth of poplar trees and that PdRanBP may regulate cell division during cell cycle progression. Taken together, our results demonstrated that PdRanBP is a nuclear, vascular tissue development-associated protein in P. deltoides.

Genes encoding Δ(8)-sphingolipid desaturase from various plants: identification, biochemical functions, and evolution.

∆(8)-sphingolipid desaturase catalyzes the C8 desaturation of a long chain base, which is the characteristic structure of various complex sphingolipids. The genes of 20 ∆(8)-sphingolipid desaturases from 12 plants were identified and functionally detected by using Saccharomyces cerevisiae system to elucidate the relationship between the biochemical function and evolution of this enzyme. Results showed that the 20 genes all can encode a functional ∆(8)-sphingolipid desaturase, which catalyzes different ratios of two products, namely, 8(Z) and 8(E)-C18-phytosphingenine. The coded enzymes could be divided into two groups on the basis of biochemical functions: ∆(8)-sphingolipid desaturase with a preference for an E-isomer product and ∆(8)-sphingolipid desaturase with a preference for a Z-isomer product. The conversion rate of the latter was generally lower than that of the former. Phylogenetic analysis revealed that the 20 desaturases could also be clustered into two groups, and this grouping is consistent with that of the biochemical functions. Thus, the biochemical function of ∆(8)-sphingolipid desaturase is correlated with its evolution. The two groups of ∆(8)-sphingolipid desaturases could arise from distinct ancestors in higher plants. However, they might have initially evolved from ∆(8)-sphingolipid desaturases in lower organisms, such as yeasts, which can produce E-isomer products only. Furthermore, almost all of the transgenic yeasts harboring ∆(8)-sphingolipid desaturase genes exhibit an improvement in aluminum tolerance. Our study provided new insights into the biochemical function and evolution of ∆(8)-sphingolipid desaturases in plants.

The Potential for Microalgae as Bioreactors to Produce Pharmaceuticals.

As photosynthetic organisms, microalgae can efficiently convert solar energy into biomass. Microalgae are currently used as an important source of valuable natural biologically active molecules, such as carotenoids, chlorophyll, long-chain polyunsaturated fatty acids, phycobiliproteins, carotenoids and enzymes. Significant advances have been achieved in microalgae biotechnology over the last decade, and the use of microalgae as bioreactors for expressing recombinant proteins is receiving increased interest. Compared with the bioreactor systems that are currently in use, microalgae may be an attractive alternative for the production of pharmaceuticals, recombinant proteins and other valuable products. Products synthesized via the genetic engineering of microalgae include vaccines, antibodies, enzymes, blood-clotting factors, immune regulators, growth factors, hormones, and other valuable products, such as the anticancer agent Taxol. In this paper, we briefly compare the currently used bioreactor systems, summarize the progress in genetic engineering of microalgae, and discuss the potential for microalgae as bioreactors to produce pharmaceuticals.

Identification of the substrate recognition region in the Δ6-fatty acid and Δ8-sphingolipid desaturase by fusion mutagenesis.

Δ8-sphingolipid desaturase and Δ6-fatty acid desaturase share high protein sequence identity. Thus, it has been hypothesized that Δ6-fatty acid desaturase is derived from Δ8-sphingolipid desaturase; however, there is no direct proof. The substrate recognition regions of Δ6-fatty acid desaturase and Δ8-sphingolipid desaturase, which aid in understanding the evolution of these two enzymes, have not been reported. A blackcurrant Δ6-fatty acid desaturase and a Δ8-sphingolipid desaturase gene, RnD6C and RnD8A, respectively, share more than 80 % identity in their coding protein sequences. In this study, a set of fusion genes of RnD6C and RnD8A were constructed and expressed in yeast. The Δ6- and Δ8-desaturase activities of the fusion proteins were characterized. Our results indicated that (1) the exchange of the C-terminal 172 amino acid residues can lead to a significant decrease in both desaturase activities; (2) amino acid residues 114-174, 206-257, and 258-276 played important roles in Δ6-substrate recognition, and the last two regions were crucial for Δ8-substrate recognition; and (3) amino acid residues 114-276 of Δ6-fatty acid desaturase contained the substrate recognition site(s) responsible for discrimination between ceramide (a substrate of Δ8-sphingolipid desaturase) and acyl-PC (a substrate of Δ6-fatty acid desaturase). Substituting the amino acid residues 114-276 of RnD8A with those of RnD6C resulted in a gain of Δ6-desaturase activity in the fusion protein but a loss in Δ8-sphingolipid desaturase activity. In conclusion, several regions important for the substrate recognition of Δ8-sphingolipid desaturase and Δ6-fatty acid desaturase were identified, which provide clues in understanding the relationship between the structure and function in desaturases.

Transcriptome sequencing of transgenic poplar (Populus × euramericana 'Guariento') expressing multiple resistance genes.

BACKGROUND: Transgenic poplar (Populus × euramericana 'Guariento') plants harboring five exogenous, stress-related genes exhibit increased tolerance to multiple stresses including drought, salt, waterlogging, and insect feeding, but the complex mechanisms underlying stress tolerance in these plants have not been elucidated. Here, we analyzed the differences in the transcriptomes of the transgenic poplar line D5-20 and the non-transgenic line D5-0 using high-throughput transcriptome sequencing techniques and elucidated the functions of the differentially expressed genes using various functional annotation methods. RESULTS: We generated 11.80 Gb of sequencing data containing 63, 430, 901 sequences, with an average length of 200 bp. The processed sequences were mapped to reference genome sequences of Populus trichocarpa. An average of 62.30% and 61.48% sequences could be aligned with the reference genomes for D5-20 and D5-0, respectively. We detected 11,352 (D5-20) and 11,372 expressed genes (D5-0), 7,624 (56.61%; D5-20) and 7,453 (65.54%; D5-0) of which could be functionally annotated. A total of 782 differentially expressed genes in D5-20 were identified compared with D5-0, including 628 up-regulated and 154 down-regulated genes. In addition, 196 genes with putative functions related to stress responses were also annotated. Gene Ontology (GO) analysis revealed that 346 differentially expressed genes are mainly involved in 67 biological functions, such as DNA binding and nucleus. KEGG annotation revealed that 36 genes (21 up-regulated and 15 down-regulated) were enriched in 51 biological pathways, 9 of which are linked to glucose metabolism. KOG functional classification revealed that 475 genes were enriched in 23 types of KOG functions. CONCLUSION: These results suggest that the transferred exogenous genes altered the expression of stress (biotic and abiotic) response genes, which were distributed in different metabolic pathways and were linked to some extent. Our results provide a theoretic basis for investigating the functional mechanisms of exogenous genes in transgenic plants.

DOP-PCR based painting of rye chromosomes in a wheat background.

To determine the appropriateness of chromosome painting for identifying genomic elements in rye, we microdissected the 1R and 1RS chromosomes from rye (Secale cereale L. var. King II) and wheat-rye addition line 1RS, respectively. Degenerate oligonucleotide primed - polymerase chain reaction (DOP-PCR) amplification of 1R and 1RS products from dissected chromosomes were used as probes to hybridize to metaphase chromosomes of rye, wheat-rye addition lines 1R and 1RS, translocation line 1RS.1BL, and allohexaploid triticale. The results showed that (i) the hybridization signal distribution patterns on rye chromosomes using 1R-derived DOP-PCR products as the probe were similar to those using 1RS-derived DOP-PCR products as the probe; (ii) 1R and (or) 1RS could not be distinguished from other rye chromosomes solely by the hybridization patterns using 1R- and (or) 1RS-derived DOP-PCR products as the probe; (iii) rye chromosomes and (or) rye chromosome fragments could be clearly identified in wheat-rye hybrids using either 1R- or 1RS-derived DOP-PCR products as the probe and could be more accurate in the nontelomeric region than using genomic in situ hybridization (GISH). Our results suggested that 1R- and (or) 1RS-derived DOP-PCR products contain many repetitive DNA sequences, are similar on different rye chromosomes, are R-genome specific, and can be used to identify rye chromosomes and chromosome fragments in wheat-rye hybrids. Our research widens the application range of chromosome painting in plants.

A Novel Strain of Bacillus cereus with a Strong Antagonistic Effect Specific to Sclerotinia and Its Genomic and Transcriptomic Analysis.

Sclerotinia, which is caused by Sclerotinia sclerotiorum, is a severe disease of oilseed rape, which is an important oil crop worldwide. In this study, we isolated a novel strain of Bacillus cereus, named B. cereus HF10, from the rhizosphere soil of the reed on the seaside of Yagzhou Bay, Sanya city, Hainan Province, China. HF10 exhibited a significant antagonistic effect on Sclerotinia sclerotiorum, with an inhibition rate of 79%, and to other species in Sclerotinia, but no antagonistic effect was found on various other fungi or bacteria. HF10 had an 82.3% inhibitory effect on the S. sclerotiorum infection of oilseed rape leaves and a 71.7% control effect on Sclerotinia infection in oilseed rape based on in vitro and in vivo experiments, respectively. The genomics and transcriptomics of HF10 and its loss of the antifungal function mutant Y11 were analyzed, and the results provided insight into potential antifungal substances. Our work provides a novel strain, HF10, for developing a promising biological control agent against Sclerotinia, which infects oilseed rape and other plants.

Natural variation in SSW1 coordinates seed growth and nitrogen use efficiency in Arabidopsis.

Seed size is controlled not only by intrinsic genetic factors but also by external environmental signals. Here, we report a major quantitative trait locus (QTL) gene for seed size and weight on chromosome 1 (SSW1) in Arabidopsis, and we found SSW1 acts maternally to positively regulate seed size. Natural variation in SSW1 contains three types of alleles. The SSW1Cvi allele produces larger seeds with more amino acid and storage protein contents than the SSW1Ler allele. SSW1Cvi displays higher capacity for amino acid transport than SSW1Ler due to the differences in transport efficiency. Under low nitrogen supply, the SSW1Cvi allele exhibits increased seed yield and nitrogen use efficiency (NUE). Locations of natural variation alleles of SSW1 are associated with local soil nitrogen contents, suggesting that SSW1 might contribute to geographical adaptation in Arabidopsis. Thus, our findings reveal a mechanism that coordinates seed growth and NUE, suggesting a potential target for improving seed yield and NUE in crops.

The role of C-terminal amino acid residues of a Δ6-fatty acid desaturase from blackcurrant.

Δ6-fatty acid desaturase is an important enzyme in the catalytic synthesis of polyunsaturated fatty acids. Using domain swapping and a site-directed mutagenesis strategy, we found that the region of the C-terminal 67 amino acid residues of Δ6-fatty acid desaturase RnD6C from blackcurrant was essential for its catalytic activity and that seven different residues between RnD6C and RnD8A in that region were involved in the desaturase activity. Compared with RnD6C, the activity of the following mutations, V394A, K395I, F411L, S436P, VK3945AI and IS4356VP, was significantly decreased, whereas the activity of I417T was significantly increased. The amino acids N, T and Y in the last four residues also play a certain role in the desaturase activity.