SrUGT76G1, the most well-studied diterpene glycosyltransferase in Stevia rebaudiana, is key to the biosynthesis of economically important steviol glycosides (SGs).  However, the molecular regulatory mechanism of SrUGT76G1 has rarely been explored.  In this study, we identified a MYB transcription factor, SrMYB1, using a yeast one-hybrid screening assay.  SrMYB1 belongs to the typical R2R3-type MYB protein and is specifically localized in the nucleus with strong transactivation activity.  The transcript of SrMYB1 is predominantly accumulated in flowers, but is also present at a lower level in leaves.  Yeast one-hybrid and electrophoretic mobility shift assays verified that SrMYB1 binds directly to the MYB binding sites in the F4-3 fragment (+50–(–141)) of the SrUGT76G1 promoter.  Furthermore, we found that SrMYB1 could significantly repress the expression of SrUGT76G1 in both epidermal cells of tobacco leaves and stevia callus.  Taken together, our results demonstrate that SrMYB1 is an essential upstream regulator of SrUGT76G1 and provide novel insight into the regulatory network for the SGs metabolic pathway in S. rebaudiana.

Sugar content is a determinant of apple (Malus×domestica Borkh.) sweetness.  However, the molecular mechanism underlying sucrose accumulation in apple fruit remains elusive.  Herein, this study reported the role of the sucrose transporter MdSUT2.1 in the regulation of sucrose accumulation in apples.  The MdSUT2.1 gene encoded a protein with 612 amino acid residues that could be localized at the plasma membrane when expressed in tobacco leaf protoplasts.  MdSUT2.1 was highly expressed in fruit and was positively correlated with sucrose accumulation during apple fruit development.  Moreover, complementary growth assays in a yeast mutant validated the sucrose transport activity of MdSUT2.1.  MdSUT2.1 overexpression in apples and tomatoes resulted in significant increases in sucrose, fructose, and glucose contents compared to the wild type (WT).  Further analysis revealed that the expression levels of sugar metabolism- and transport-related genes SUSYs, NINVs, FRKs, HXKs, and TSTs increased in apples and tomatoes with MdSUT2.1 overexpression compared to WT.  Finally, unlike the tonoplast sugar transporters MdTST1 and MdTST2, the promoter of MdSUT2.1 was not induced by exogenous sugars.  These findings provide valuable insights into the molecular mechanism underlying sugar accumulation in apples.

ABA receptors (PYR/PYL/RCAR) play a central role in the water loss control of plants.  A previous report indicated that TaPYL4 is a critical gene in wheat that improves grain production under drought conditions and increases water use efficiency.  In this study, we analyzed the sequence polymorphisms and genetic effects of TaPYL4s.  Based on isolated TaPYL4 genes from chromosomes 2A, 2B and 2D, three haplotypes were detected in the promoter region of TaPYL4-2A, and two haplotypes were present in TaPYL4-2B and TaPYL4-2D, respectively.  Marker/trait association analysis indicated that TaPYL4-2A was significantly associated with plant height in 262 Chinese wheat core collection accessions, as well as the drought tolerance coefficient (DTC) for plant height in 239 wheat varieties from Shanxi Province in multiple environments.  However, the frequencies of favored drought-tolerant haplotype TaPYL4-2A-Hap2 were considerably low, accounting for only 10%, and lines with this certain Hap could be reserved in the breeding program.  TaPYL4-2B was significantly associated with grain number, and the favored haplotype TaPYL4-2B-Hap1 was the dominant allele of above 90% in the collection.  For TaPYL4-2D, there were no significant differences in these traits between the two haplotypes in either of the two panels.  These results indicate that variation might lead to functional differentiation among the homoeologs and the haplotypes had undergone artificial selection during breeding.  Two molecular markers developed to distinguish these haplotypes could be used for breeding in water-limited regions.

Mycoplasma gallisepticum HS strain (MG-HS) is a pathogen that causes chronic respiratory disease (CRD) in chicken, which is characterized by host respiratory inflammatory damage, brings huge economic losses to the poultry industry.  Recently, emerging Chinese herbal medicines (CHM) have been used to treat CRD.  This study was aimed to investigate the preventive and therapeutic effects and their potential mechanisms of Chinese herbal medicinal formulae (CHMF), which consisted of 10 kinds of Chinese herbal medicine including Scutellaria, Houttuynia cordate and licorice, on MG-induced CRD in chickens.  With respect to the preventive effect, the results showed that CHMF could effectively recover the MG-induced decrease on body weight and feed conversion ratio.  Histopathological analysis showed that both prevention and treatment of CHMF could significantly alleviate the severe respiratory inflammation induced by MG infection.  Moreover, compared with the MG infection group, both the prevention groups and the treatment groups of CHMF could effectively reduce the expression of MG adhesion protein (pMGA1.2) to inhibit the proliferation of MG, and thus effectively inhibit the expression of MG-induced inflammatory factors interleukin-1β (IL-1β) and tumour necrosis factor-α (TNF-α).  In summary, these findings confirm that CHMF can protect chickens from various tissue damage caused by MG infection and has no adverse effects on the performance of chickens in the short term.  And its efficacy against MG is equal to or better than that of tiamulin.

In 2009, an emerging citrus viral disease caused by Citrus chlorotic dwarf-associated virus (CCDaV) was discovered in Yunnan Province of China.  However, the occurrence and spread of CCDaV in other citrus-growing provinces in China is unknown to date.  To better understand the distribution and molecular diversity of CCDaV in China, a total of 1 772 citrus samples were collected from 11 major citrus-growing provinces and were tested for CCDaV by PCR.  Among these, 134 citrus samples from Guangxi, Yunnan and Guangdong were tested positive for CCDaV, demonstrating that the occurrence and spread of CCDaV are increasing in China.  The complete genome sequences of 17 CCDaV isolates from different provinces and hosts were sequenced.  Comparisons of the whole-genome sequences of the 17 CCDaV isolates as well as the 15 isolates available in GenBank revealed that the sequence identity was about 99–100%, showing that the CCDaV isolates were highly conserved.  Phylogenetic studies showed that the 32 CCDaV isolates belonged to four different groups based on geographical origins and host species, and that CCDaV isolates from China and Turkey were clustered into different groups.  The results provide important information for clarifying the distribution and genetic diversity of CCDaV in China.

This study investigated the influence of activities of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and lactate dehydrogenase-B (LDH-B) on the colour stability of mutton.  From 60 sheep (Bayannur mutton sheep), 15 longissimus dorsi (LD) muscles were selected on the basis of colour stability (R630/580 and a* value) during the storage and classified into three groups (5 for each group) as high colour stability (HCS), intermediate colour stability (ICS) and low colour stability (LCS).  The activities of GAPDH and LDH-B, muscle colour attributes, nicotinamide adenine dinucleuotide (NADH) concentration and lactate concentration were measured.  The samples in HCS had higher activities of GAPDH and LDH-B than the samples in the LCS, and the samples in the HCS group also possessed higher NADH and lower lactate concentration.  The higher activity of dehydrogenase enzyme may result in higher NADH concentrations and colour stability in muscle tissue.  The results suggest that the activity of GAPDH and LDH-B may also play a role in maintaining colour stability.

    The effect of long-term fertilization on soil denitrifying communities was analysed by measuring the abundance and diversity of the nitrous oxide (N₂O) reductase gene, nosZ.  Soil samples were collected from plots of a long-term fertilization experiment established in 1982 in Suining City, China.  The fertilizer treatments were no fertilizer (CK), three chemical fertilizer (CF) treatments (N, NP, NPK), manure (M) alone, and manure with chemical fertilizers (NM, NPM, NPKM).  The abundance and diversity of the denitrifying bacteria were assessed by real-time quantitative PCR, terminal restriction fragment length polymorphism (T-RFLP), and cloning and sequencing of nosZ genes.  The diversity and abundance of nosZ-denitrifiers was higher in soil amended with manure and chemical fertilizers (CFM) than in soil amended with CF alone, and the highest in topsoil (0–20 cm).  The nosZ-denitrifier community composition was more complex in CFM soil than in CF soil.  Specific species were detected only in the CFM soil.  The abundance of nosZ-denitrifier in the NPKM treatment was approximately two times higher than that in the CK, N, and NPK treatments.  Most of the cloned nosZ sequences were closely related to nosZ sequences from Bradyrhizobiaceae and Rhodospirillaceae in Alphaproteobacteria.  Of the measured abiotic factors, soil organic matter correlated significantly with the abundance (P<0.01); available phosphorus correlated significantly with the topsoil community composition (P<0.01), whereas soil organic matter correlated significantly with the subsoil (20–90 cm) community composition (P<0.01). This study demonstrated that long-term CFM fertilization affected both the abundance and composition of the nosZ-denitrifier community. 

In biological research, chicken embryos are a classic experimental model for the exploration of the embryonic development and cell differentiation.  Transferring exogenous substances into chicken embryos for producing medical antibodies has been widely used in the production practice.  However, there are few studies about the effect of the different injection site and dosage on chicken embryos.  The aim of this study was to explore the effects of different injection sites and dosages on chicken embryo hatching rate and development, so as to provide a basis for further studies using the chicken embryo model.  Freshly laid eggs (Rugao yellow chicken) were injected with different doses of saline at the tip, equatorial plane and the blunt end of the egg shell, respectively.  Egg hatching rate was recorded and compared among injection sites and different doses.  A trypan blue stain was also injected at the aforementioned sites and the growth of chicken embryos was observed.  The SPSS (statistical package for the social science) software was used to analyze the relationship between the chicken eggs hatching rate and the different injection sites or the different dosages.  The experimental results showed that there were significant differences on egg hatching rates among the different injection sites and doses (P<0.05).  The hatchability of the blunt end injection group was significantly higher than that of the other two sites.  The egg hatching rate decreased with increased saline doses.  The egg hatching rate of the 100 µL saline injection group was higher than the 200 and 300 µL dosage groups.  Ultimately, we suggest that the optimal chicken embryo injection process is during early development, at the blunt end site with a dose less than 100 µL to minimize damage to the egg.

Plant cell walls constitute the skeletal structures of plant bodies, and thus confer lodging resistance for grain crops.  While the basic cell wall synthesis machinery is relatively well established now, our understanding of how the process is regulated remains limited and fragmented.  In this study, we report the identification and characterization of the novel rice (Oryza sativa L.) brittle culm16 (brittle node; bc16) mutant.  The brittle node phenotype of the bc16 mutant appears exclusively at nodes, and resembles the previously reported bc5 mutant.  Combined histochemical staining and electron microscopy assays revealed that in the bc16 mutant, the secondary cell wall formation and thickening of node sclerenchyma tissues are seriously affected after heading.  Furthermore, cell wall composition assays revealed that the bc16 mutation led to a significant reduction in cellulose and lignin contents.  Using a map-based cloning approach, the bc16 locus is mapped to an approximately 1.7-Mb region of chromosome 4.  Together, our findings strengthen evidence for discretely spatial differences in the secondary cell wall formation within plant bodies.

A total of 50 endophytic bacterial isolates were obtained from Kobreasia capillifolia at alpine grasslands in the Eastern Qilian Mountains on the Tibetan Plateau in China.  Based on the sequencing and phylogenetic analysis of 16S rDNA genes, all isolates phylogenetically related closely to Bacillus, Acinetobacter, Stenotrophomonas, Brevundimonas, Arthrobacter, Curtobacterium, Paenibacillus, Plantibacter, Promicromonospora, Serratia, and Microbacterium, among which Bacillus was the predominant genus (47.8% of the total number of endophytic isolates).  These isolates possessed different biological functions.  In 50 endophytic bacteria, 42 isolates produced indole acetic acid (IAA) on King medium.  There were seven isolates showing potency of mineral phosphate solubilization in Pikovaskaia’s (PKO) liquid medium.  Seven isolates exhibited antagonistic effect against Fusarium avenaceum, Colletotrichum coccodes and Phoma foveata.  This was the first report on diversity and plant growth promotion of endophytic bacteria from K. capillifolia on alpine grassland in the Eastern Qilian Mountains, Chain.  It is essential for revealing the relationship among plant, microorganism, and the special environment because the potential function of endophytic bacteria made a contribution to the interactions of plants and endophytic bacteria.

    In this study, two ploidy mutant lines of Myrica rubra cv. Dongkui (DK) were identified and named as DB1 and DB2. The floral organ, leaf cell structure, ploidy, and number of chromosomes of the two mutants were investigated. Meanwhile, anthocyanin contents at different developmental stages were analyzed, and the Cy-3-glu contents of DB1 and DB2 at the full ripe stages are significantly higher than that of DK by 27.84 and 23.51%, respectively. Furthermore, 6 RNA libraries at two developmental stages (young fruit stage and full ripe stage) were built for RNA-Seq. By mapping to the reference database, 28 407, 28 043, and 28 683 genes were detected in the young fruit of DB1, DB2, and DK, respectively, while 28 040, 22 256, and 27 351 genes were detected in the full ripe stage, respectively. There were 281 differentially expressed genes between DB1 and DK, with 123 and 158 genes up-regulated and down-regulated, respectively, and 47 differentially expressed genes between DB2 and DK, of which 8 and 39 genes were up-regulated and down-regulated. Using real-time PCR, the expression levels of the eight functional genes at different developmental stages of the fruit were also analyzed. These comprehensive analyses showed that both mutants are different from DK, which is the result of natural doubling of ploidy, thereby generating a pleiotropic effect. As we known, it is the first report to study the relationship between bayberry ploidy alterations and genes involved in regulation of fruit mutations, which will help to identify the morphological and cytological characteristics of M. rubra germplasm, and provide a theoretical basis and technical support for genetic improvement and creation of breeding resources.

    As selenium and carotenoids are essential micronutrients, the determination of their concentrations in different varieties is important in the breeding of foxtail millet (Setaria italica L. P. Beauv.). To identify selenium- and carotenoid-enriched foxtail millet varieties and to analyze correlations between trace elements and agronomic traits, we measured the selenium and carotenoid concentrations of 200 Chinese accessions by high-performance liquid chromatography and atomic fluorescence spectrometry. Our analysis revealed that lutein concentration in 200 foxtail millet accessions followed normal distribution and average was 3.1 μg g^–1. The mean value of zeaxanthin concentration in 200 accessions was 8.6 μg g^–1. Lutein and zeaxanthin concentrations were higher in the foxtail millet from Liaoning than in varieties from other locations, with averages of 10.0 and 3.5 μg g^–1, respectively. The average measured selenium concentration was 100.3 μg kg^–1. The highest average selenium concentration, 110.3 μg kg^–1, was found in varieties from Shanxi. Varieties from Inner Mongolia had the lowest average selenium concentration, 84.7 μg kg^–1, which was significantly lower (P<0.05) than that of Shanxi. Selenium concentrations of 23 varieties were higher than 117.9 μg kg^–1, accounting for 11.5% of the total, thereby were considered to be enriched in selenium. In addition, we identified 29 lutein-enriched varieties (>4.27 μg g^–1) and 30 zeaxanthin-enriched ones (>12.63 μg g^–1), which corresponded to 14.5 and 15% of tested accessions, respectively. Correlation analysis revealed that selenium concentration was significantly positively correlated with spikelet length (P<0.01), while zeaxanthin concentration was significantly correlated with grass weight (P<0.05) and spikelet length (P<0.01). No correlation was found between lutein concentration and agronomic characters, selenium content or zeaxanthin content. Our results should contribute substantially to the selection of suitable varieties for the development of plants with desired levels of these nutritionally important elements. These results will significantly contribute towards selection of the most suitable varieties for obtaining plants with desired levels of these nutritionally important elements.

  Controlling the height of crops plays a crucial role for their yields. The large scale utilization of semi-dwarf varieties has greatly improved crop yield, providing an effective support for world food security. In rice, a main food for over half of the world’s population, a number of dwarf loci have been identified. However, most of them are recessive, such as the ‘green revolution’ gene sd1. To gain more beneficial loci for rice breeding programs, exploring new mutations is needed, especially the dominant loci which can be used broadly for hybrid breeding. Here, we isolated a novel dominant dwarf rice mutant, slr1-d5. All of the internodes of slr1-d5 are reduced. We find that the responsiveness of slr1-d5 to gibberellin (GA), GA₃, was significantly reduced. Map-based cloning revealed that the dominant dwarfism of slr1-d5 was caused by an amino acid substitution in the N-terminal TVHYNP domain of rice DELLA protein, SLR1, where the conserved amino acid Pro (P) was substituted to His (H). Our findings not only further prove the pivotal role of TVHYNP motif in regulating SLR1 stability, but also provide a new dwarf source for improvement of rice germplasms.

Direct-sowing establishment method has great significance in improving winter oilseed rape (Brassica napus L.) production and guaranteeing edible oil security in China. However, nutrient responses on direct sown winter oilseed rape (DOR) performance and population development dynamic are still not well understood. Therefore, five on-farm experiments were conducted in the reaches of the Yangtze River (RYR) to determine the effects of nitrogen (N), phosphorus (P), and potassium (K) deficiencies on population density, dry matter production, nutrient uptake, seed yield, and yield components of DOR plants. Four fertilization treatments included the balanced NPK application treatment (NPK, 180 kg N, 39.3 kg P, 100 kg K, and 1.8 kg borax ha–1) and three nutrient deficiency treatments based on the NPK treatment, i.e., –N, –P, and –K. The results indicated that DOR population density declined gradually throughout the growing season, especially at over-wintering and pod-development stages. Nutrient deficiency decreased nutrient concentration in DOR plants, limited dry matter production and nutrient uptake, and thereby exacerbated density reduction during plants growth. The poor individual growth and reduced population density together decreased seed yield in the nutrient deficiency treatment. Averaged across all the experiments, seed yield reduced 61% by N deficiency, 38.3% by P deficiency, and 14.4% by K deficiency. The negative effects of nutrient deficiency on DOR performances followed the order of –N>–P>–K, and the effects were various among different nutrient deficiencies. Although N deficiency improved DOR emergence, but it seriously limited dry matter production and nutrient uptake, which in turn led to substantial plants death and therefore resulted in a very low harvested density. The P deficiency significantly reduced initial density, limited plants growth, and exacerbated density reduction. The K deficiency mainly decreased individual growth and yield, but did not affect density dynamic. Our results highlighted the importance of balanced NPK application in DOR production, suggesting that management strategy of these nutrients should be comprehensively considered with an aim to build an appropriate population structure with balanced plant density and individual growth.

In order to generate an antibody against a small hapten molecule, the hapten is cross-linked with carrier protein to make it immunogenic. In this study, the hapten (ochratoxin A, OTA) was coupled to ovalbumin (OVA) by an active ester reaction. To develop a technique for detecting the conjugation, the hapten-protein conjugate (OTA-OVA) was characterized thoroughly by immunoarray technology, ultraviolet (UV) spectroscopy and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), respectively. The molecular weight of OTA-OVA was 50 350.141 Da, and the molecular weight of OVA was 44 887.506 Da, which were determined by MALDI-TOF-MS, respectively. In OTA-OVA, the molecular coupling ratio was 13:1 by MALDI-TOF-MS while the molecular coupling ratio was 10:1 by UV. In this experiment, UV and MALDI-TOF-MS were selected as the efficient methods to evaluate the cross-linking effect and calculate the molecular coupling ratio.

Magnaporthe oryzae, a filamentous ascomycete fungus, is well known as the causal agent of rice blast. With thetechnology of suppression subtractive hybridization (SSH), it was previously found that MGG_06001 (or named MoNEM1),a gene of M. oryzae homologous to the NEM1 (nuclear envelope morphology protein 1) gene of baker’s yeast(Saccharomyces cerevisiae), is differentially expressed between the mature appressium and the conidium and mycelium.This study aimed to characterize the function of MoNEM1 gene by knocking it out using the method of target genereplacement. The ΔMonem1 mutants exhibited reduced mycelial growth and conidiation. However, disruption of MoNEM1gene does not affect the pathogenicity of M. oryzae on barley and rice.

Mycoplasma gallisepticum (MG) is a common avian pathogen that mainly infects poultry, causing significant reductions in body weight gain and egg production, along with damage to immune organs and immunosuppression. MG is susceptible to co-infections with other pathogens, leading to increased mortality rates and significant economic losses in the global poultry industry. While antibiotics have been extensively applied worldwide to treat MG infections in poultry production, concerns regarding antibiotic resistance and residue remain prevalent. Traditional Chinese medicine (TCM), renowned for its natural, safe, and non-toxic properties, has shown significant anti-inflammatory and immune-enhancing effects. This study aimed to investigate the protective effect of TCM on production performance and its impact on MG-induced immunosuppression through the MAPK/ERK/JNK signaling pathway in chickens. Our results showed that TCM alleviated the negative effects of MG infection on production performance, as evidenced by improvements in body weight gain, feed conversion rate, survival rate, and immune organ index. TCM exhibited direct inhibition of the MG proliferation in vitro and in vivo. Furthermore, TCM treatment promoted the normalization of trachea and lung tissue structure in MG-infected chickens, leading to a significant reduction in inflammatory damage. Moreover, following the treatment with the TCM, the production of pro-inflammatory cytokines Interleukin-1β (IL-1β) and Tumor Necrosis Factor-α (TNF-α) decreased significantly, accompanied by the downregulation of pro-apoptotic genes caspase3, caspase9, and BAX, both in vitro and in vivo. A mechanism-based study showed that in vitro and in vivo treatment with the TCM significantly reduced the expression of key proteins, including early growth response gene1 (EGR1), p-ERK, p-JNK, and p-JUN. Altogether, TCM improved body weight gain, inhibited pro-inflammation responses, and alleviated tissue damage by inhibiting the MAPK/ERK/JNK signaling pathway to protect the performance and immune system of MG-infected chickens.