Duck hepatitis B virus (DHBV) shares many basic characteristics with hepatitis B virus (HBV) and is an attractive model for vaccine development.  In this study, DHBV DNA vaccines were designed to express envelope and capsid fusion proteins to enhance the breadth of immune response in ducks.  Attenuated Salmonella typhimurium (SL7207) was used as a carrier and adjuvant to boost the magnitude of immune response.  Based on this strategy, novel DNA vaccines (SL7207-pVAX1-LC and SL7207-pVAX1-SC) were generated.  Growth kinetics, genetic stabilities and relative transcription levels of the L, S and C genes introduced by these vaccine strains were measured before inoculation to guarantee safety and efficacy.  The relative transcript levels of the CD4 and CD8 T genes and the antibody levels (IgY) in ducks receiving the vaccines were higher than those in single gene delivered groups.  Additionally, the copy number of covalently closed circular DNA in hepatocytes after DHBV challenge also provided evidence that our fusion vaccines could enhance the protective efficiency against DHBV infection in ducks.

    The capsid (Cap) protein, which is the only structural protein of duck circovirus (DuCV), is the most important antigen for the development of vaccines against DuCV and the virus’s serological diagnostic methods. In order to use yeast expression system to produce a large quantities of DuCV Cap protein which is close to its natural form to display the antigen peptides perfectly, the Cap gene was optimized into the codon-optimized capsid (Opt-Cap) gene towards the preference of yeast firstly. Then, the genes of Cap and Opt-Cap were separately cloned into pPIC9K plasmid and transformed into Picha pastoris GS115. The strains that displayed the phenotype of Mut+ and contained multiple inserts of expression cassette were selected from those colonies. After the induction expression, the secretory type of Cap protein, which was about 43 kDa, was best expressed under 0.5% (v/v) methanol and sorbitol induction. Compared with the Cap gene, the expression level of Opt-Cap gene was much higher. What’s more, the purified Cap protein had a good reactivity to its specific polyclone antibody and DuCV-positive serum, and it was able to self-assemble into virus-like particles (VLPs). These VLPs, with a diameter of 15–20 nm and without a nucleic acid structure, showed a high level of similarity to DuCV particles in size and shape. All of the results demonstrated that, based on the codon-optimization, it is suitable to use the P. pastoris expression system to produce DuCV VLPs on a large scale. It is the first time that a large amounts of DuCV VLPs were produced successfully in P. pastoris, which might be particularly useful for the further studies of serological diagnosis and vaccines of DuCV.

Understanding the response of soil respiration to global warming in agro-ecosystem is crucial for simulating terrestrial carbon (C) cycle.  We conducted an infrared warming experiment under conventional tillage (CT) and no-tillage (NT) farmland for winter wheat and summer maize rotation system in North China Plain (NCP).  Treatments include CT with and without warming (CTW and CTN), NT with and without warming (NTW and NTN).  The results indicated that warming had no significant effect on soil moisture in irrigated farmland of NCP (P>0.05).  The elevated average soil temperature of 1.1–1.6°C in crop growing periods could increase annual soil CO₂ emission by 10.3% in CT filed (P>0.05), but significantly increase it by 12.7% in NT field (P<0.05), respectively.  The disturbances such as plowing, irrigation and precipitation resulted in the obvious soil CO₂ emission peaks, which contributed 36.6–40.8% of annual soil cumulative CO₂ emission.  Warming would enhance these soil CO₂ emission peaks; it might be associated with the warming-induced increase of autotrophic respiration and heterotrophic respiration.  Compared with un-warming treatments, dissolved organic carbon (DOC) and soil microbial biomass carbon (MBC) in warming treatments were significantly increased by 11.6–23.4 and 12.9–23.6%, respectively, indicating that the positive responses of DOC and MBC to warming in both of two tillage systems.  Our study highlights that climate warming may have positive effects on soil C release in NCP in association with response of labile C substrate to warming.

Potassium (K) deficiency is one of the major abiotic stresses which has drastically influenced maize growth and yield around the world. However, the physiological mechanism of K deficiency tolerance is not yet fully understood. To identify the differences of root morphology, physiology and endogenous hormones at different growing stages, two maize inbred lines 90-21-3 (tolerance to K deficiency) and D937 (sensitive to K deficiency) were cultivated in the long-term K fertilizer experimental pool under high potassium (+K) and low potassium (–K) treatments. The results indicated that the root length, volume and surface area of 90-21-3 were significantly higher than those of D937 under –K treatment at different growing stages. It was noteworthy that the lateral roots of 90-21-3 were dramatically higher than those of D937 at tasselling and flowering stage under –K treatment. Meanwhile, the values of superoxide dismutase (SOD) and oxidizing force of 90-21-3 were apparently higher than those of D937, whereas malondialdehyde (MDA) content of D937 was obviously increased. Compared with +K treatment, the indole-3-acetic acid (IAA) content of 90-21-3 was largely increased under –K treatment, whereas it was sharply decreased in D937. On the contrary, abscisic acid (ABA) content of 90-21-3 was slightly increased, but that of D937 was significantly increased. The zeatin riboside (ZR) content of 90-21-3 was significantly decreased, while that of D937 was relatively increased. These results indicated that the endogenous hormones were stimulated in 90-21-3 to adjust lateral root development and to maintain the physiology function thereby alleviating K deficiency.

MicroRNAs (miRNAs) are small, single stranded, non-coding RNA molecules, about 19–25 nucleotides in length, which regulate the development and functions of reproductive system of mammal. To discover novel miRNAs and identify the differential expression of them in ovine ovary and testis tissues, the study constructed two libraries by using next-generation sequencing technologies (Solexa high-throughput sequencing technique). As a result, 9 321 775 and 9 511 538 clean reads were obtained from the ovary and testis separately, which included 130 562 (90 genes of ovary) and 56 272 (85 genes of testis) of known miRNAs and 486 potential novel miRNAs reads. In this study, a total of 65 conserved miRNAs were significantly differentially expressed (P<0.01) between the two samples. Among them, 28 miRNAs were up-regulated and 3 miRNAs were down-regulated on ovary compared with testis. In addition, the known miRNAs with the highest expression level (5 miRNAs) and 30 novel miRNAs with the functions related to reproduction were validated using the real-time quantitative RT-PCR. Moreover, the gene ontology (GO) annotation and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis showed that differentially expressed miRNAs were involved in ovary and testis physiology, including signal transduction, gonad development, sex differentiation, gematogenesis, fertilization and embryo development. The results will be helpful to facilitate studies on the regulation of miRNAs during ruminant reproduction.

Potassium is an important nutrient element requiring high concentration for photosynthetic metabolism. The potassium deficiency in soil could inhibit soybean (Glycine max (L.) Merr.) photosynthesis and result in yield reduction. Research on the photosynthetic variations of the different tolerant soyben varieties should provide important information for high yield tolerant soybean breeding program. Two representative soybean varieties Tiefeng 40 (tolerance to K+ deficiency) and GD8521 (sensitive to K+ deficiency) were hydroponically grown to measure the photosynthesis, chlorophyll fluorescence parameters and Rubisco activity under different potassium conditions. With the K-deficiency stress time extending, the net photosynthetic rate (Pn), transpiration rate (Tr) and stomatal conductance (Gs) of GD8521 were significantly decreased under K-deficiency condition, whereas the intercellular CO2 concentration (Ci) was significantly increased. As a contrast, the variations of Tiefeng 40 were almost little under K-deficiency condition, which indicated tolerance to K+ deficiency variety could maintain higher efficient photosynthesis. On the 25th d after treatment, the minimal fluorescence (F0) of GD8521 was significantly increased and the maximal fluorescence (Fm), the maximum quantum efficiency of PSII photochemistry (Fv/ Fm), actual photochemical efficiency of PSII (ΦPSII), photochemical quenching (qP), and electron transport rate of PSII (ETR) were significantly decreased under K+ deficiency condition. In addition, the Rubisco content of GD8521 was significantly decreased in leaves. It is particularly noteworthy that the chlorophyll fluorescence parameters and Rubisco content of Tiefeng 40 were unaffected under K+ deficiency condition. On the other hand, the non-photochemical quenching (qN) of Tiefeng 40 was significantly increased. The dry matter weight of Tiefeng 40 was little affected under K+ deficiency condition. Results indicated that Tiefeng 40 could avoid or relieve the destruction of PSII caused by exceeded absorbed solar energy under K-deficiency condition and maintain natural photosynthesis and plant growth. It was an essential physiological mechanism for low-K-tolerant soybean under K-deficiency stress.

This study is first to investigate proteomic changes in sheep sperm induced by carbon ion radiation using two-dimensional electrophoresis (2-DE) analysis in the project of breeding a new variety of sheep. Differential expression proteins were detected using the PDQuest 8.0 software after staining with Coomassie blue. Valid spots were then analyzed through liquid chromatography tandem mass spectrometry (LC-MS/MS). Among the 480 total protein spots displayed in 2-D gels, 6 specific protein spots were observed in sperm gels. A search against protein sequences in the National Center for Biotechnology Information databases (NCBI) indicated that differentially expressed proteins correspond to two proteins, identified to be enolase and transcription factor AP-2-alpha (TFAP-2α). The two proteins were up-regulated in the irradiated sperm. To the best of our knowledge, this study is the first to identify proteomic changes induced by carbon ion radiation in sheep sperm. The analysis of differential expression protein may be useful in identifying new breeding markers in sheep reproduction and in clarifying the mechanisms involved in irradiation or space breeding.

To further research the regulatory network of pyruvate dehydrogenase kinase (designated as TaPDK) in physiological male-sterility (PHYMS) of wheat induced by chemical hybridizing agent (CHA) SQ-1, an anther cDNA library was constructed, and the proteins interacting with TaPDK were screened via yeast two-hybrid technique. Subsequently, a few candidate proteins in nucleotide expression levels were detected by real-time quantitative PCR. Yeast-two hybrid screening was performed by mating yeast strain Y2HGold containing BD-TaPDK bait plasmid with yeast strain Y187 including anther cDNA library plasmid. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Ade/ -His/-Leu/-Trp) (QDO), and further were incubated on QDO medium containing AbA and X-α-Gal. The interactions between TaPDK and the proteins obtained from positive colonies were further confirmed by co-transformation validation. After plasmids DNA were extracted from blue colonies and sequenced, the sequences results were analyzed by bioinformatic methods. Finally, 24 colonies were obtained, including eight genes, namely non-specific lipid-transfer protein precursor (TanLTP), polyubiquitin (TaPUbi), glyceraldehyde-3-phosphate dehydrogenase, proliferating cell nuclear antigen (TaPCNA), CBS domain containing protein (TaCBS), actin, guanine nucleotide-binding protein beta subunit, chalcone synthase, and three new genes with unknown function. The results of quantitative RT-PCR showed that the expression levels of TanLTP, TaPUbi, and TaPCNA were obviously up-regulated in PHYMS anther, and TaCBS expression was only increased at the tricellular stage in PHYMS anther compared with in fertile lines. Whereas, the expression of TaPDK was obviously down-regulated in PHYMS lines. Collectively, these datas indicated that the majority of candidate proteins might be related to pollen abortion in PHYMS lines, which further suggested that TaPDK plays multiple roles in pollen development, besides participating in regulating pyruvate dehydrogenase complex activity.

Two genetic linkage maps, constructed by DH and RILs populations derived from the same parents, were carried out for the identification and comparison of QTLs controlling yield traits across different years in rice (Oryza sativa L.). A total of 194 SSR and STS markers were used in two maps, of which 114 markers were same. The distribution of Samgang allele was higher in RILs population than it in DH population. Comparing with DH population, RILs population has more lines with higher yield and wider phenotypic transgressive segression for yield traits. Although most of QTLs for the same trait were different in two populations across different years, 8 QTLs (including gwp11.1, spp5.1, spp10.1, spp11.2, ssr1.1, ssr11.1, tgw9.1 and tgw11.1) were detected over 2 yr. It is important to note that ppp10.1, spp10.1 and tgw9.1 were identified in two populations, while spp10.1 and tgw9.1 were simultaneity observed across different years. Epistatic effects were more important than additive effects for PPP, SPP, yield in DH population and TGW, yield in RILs population. Epistatic effects of DH and RILs populations were different on the same genetic background in the present study, which illuminated the QE interaction played an important role on epistatic effect. Identification and comparison of QTLs for yield traits in DH and RILs populations should provide various and more precise information. The QTLs identified in present study would be valuable in marker-assisted selection program for improving rice yield.

Mitogen activated protein kinases (MAPK) cascades based on protein phosphorylation play an important role in plant growth and development. In this study, we have identified 15 putative members of the wheat MAPK gene (TaMPK) family through an in silico search of wheat expressed sequence tags (EST) databases based on the presence of amino acid sequence of Arabidopsis and rice MAPKs. Phylogenetic analyses of MAPKs from wheat, rice and Arabidopsis genomes have classified them into seven subgroups (A, B, C, D, E, F, and G). Using the available EST information as a source of expression data, the MAPK family genes from Triticum aestivum were detected in diverse tissues. Further expression analysis of the MAPKs in NCBI EST database revealed that their transcripts were most abundant in callus (20%), followed by leaf (12%) and inflorescence (12%). Most MAPK family genes showed some tissue specificity.

This experiments were conducted in Nanjing (118º50´E, 32º02´N) and Xuzhou (117°11´E, 34°15´N), Jiangsu Province, China, to study the response of fiber quality to the N concentration of the leaf subtending boll in cotton (Gossypium hirsutum L.). Results suggested that the N dilution curve of the leaf subtending boll can accurately indicate the stagespecific plant N status for fiber development. Fiber strength is likely to be the most variable fiber quality index responding to the leaf N variation which is different in cultivars. Fiber length was the most stable index among strength, length, micronaire, and elongation. There existed an optimum leaf N concentration for fiber strength development in each stage. The optimum leaf N regression curve was very close to the N dilution curve in the middle positional fruiting branches under the 240 kg N ha-1 soil N application rate.

Oryza officinalis is one of the important wild species in the tertiary gene pool of Oryza sativa. It has a number of elite genes for rice breeding in resistance or tolerance. However, breeding barriers are so serious that the gene transfer is much difficult by sexual cross method between O. sativa and O. officinalis. Characteristics of the breeding barriers were systemically studied in this paper. When both the diploid (AA, 2n=2x=24) and autotetraploid (AAAA, 2n=4x=48) cultivated rice were crossed as maternal parents with O. officinalis (CC, 2n=2x=24), none F1 hybrid seeds were obtained. The young hybrid ovaries aborted at 13-16 d after pollinations (DAP). By rescuing hybrid embryos, in vitro F1 plantlets were obtained in 2x×2x combinations with the crossabilities lower than 0.5%. Lower rates of double-fertilization and abnormal development of hybrid embryo and endosperm were mainly observed in both combinations of 2x×2x and 4x×2x. Free endosperm nuclei in hybrid degenerated early at 1 DAP in a large scale. Almost no normal endosperm cells formed at 3 DAP. Development of a lot of embryos ceased at globular- or pear-shaped stage as well as some degenerated gradually. The hybrid plantlets were both male and female sterility. Due to the abnormal development, a diversity of abnormal embryo sacs formed in hybrids, and hybrid pollen grains were typically abortive. It showed that conflicts of genome A and C in hybrid induced abnormal meioses of meiocytes.