The "three-line" hybrid breeding method is one of the important ways to utilize crop heterosis. When the hybrid combinations were prepared, it was found that although some male-sterile lines performed well agronomically, the fertility in their F₁ generation could not be fully restored. Therefore, it was speculated that there may be a gene encoding for a suppressor for the restoration of male sterility. The speculated suppressor coding gene was named as Rf-I, an inhibitor of fertility restorer gene. In this study, the RN-type cytoplasmic male sterility (CMS) soybean (Glycine max) line JLCMS89A and restorer line JLR92 were used as genetic materials. First, the genetic analysis was performed by using F₁ and backcross (BC₁) generation populations. Then, bulk segregant analysis (BSA) high-throughput sequencing was combined with simple sequence repeat (SSR) molecular marker technology was used to preliminarily map the Rf-I gene. The results showed that the restoration suppression trait was controlled by a single dominant gene located in the nuclear genome of the sterile line. The Rf-I was initially mapped between 38.387 and 39.890 Mb at the Chromosome 9, within the range of about 1.503 Mb. This study provides theoretical and experimental basis for subsequent research on the fine mapping of Rf-I gene and characterization of its function. In addition, the result can guide the choice of the RN-type soybean cytoplasmic male sterile lines to make hybrid soybean.

To protect and rationally utilize Russula resources and investigate the regulation of protective picking on the structure of ectomycorrhizal fungi in Fagaceae symbiotic host of mushroom Russula could provide a theoretical basis to improve Russula yield increasing in Natural Russula forest. In present study, using morphology analysis and ribosomal DNA internal transcribed spacer (rDNA ITS) sequencing, the dominant species was identified as Russula griseocarnosa in Fujian, while in some part of eastern Fujian, the major species was identified as R. rosea. In addition, the combination of morphology and rDNA ITS sequencing were used to compare the differences in the composition and abundence of ectomycorrhizal communities in Fagaceae symbiotic tree caused by non-protective and protective picking (that is, the last stubble of the previous year (about late August) all picked and not picked) in a Russula nature reserve area in Jianou, Fujian. A total of 34 OTUs (operational taxonomic units) was identified in ectomycorrhizal fungi from root tips of Fagaceae. Among them, 4 species, 9 genus with unknown species, 3 family with unknown genus and 1 order with unknown family were identified. The core communities were Lactarius, Thelephoraceae, Russula and Cortinarius. The protective picking of Russula in the last crop increased the abundance of Russula and Thelephoraceae in the second year, while the abundance of Lactarius decreased. The present study clarified the types of mycorrhizal fungi and found different harvesting of Russula in the last stubble could affect the symbiosis rate of Russula with the Fagaceae and flora structure of Fagaceae roots in the next year, which could provide a reference for the artificial promotion of Russula production as well as restoration and transformation of the Russula protected forest ecosystem.

The ubiquitin-proteasome pathway is widely involved in various stages of plant growth and development. The ubiquitin-conjugating enzyme E2 is the key enzyme in the pathway of enzyme-catalyzed and plays an important role in process of substrate ubiquitination. In this study, the StUBC12 (GenBank No. XM_006343293.2) was cloned from the potato (Solanum tuberosun) cultivar 'Atlantic' leaf cDNA. The full-length of the gene was 2 112 bp, and the CDS region is 444 bp. It was a split gene containing 3 introns and 4 exons. Bioinformatics analysis of StUBC12 indicated that the gene was a non-transmembrane protein encoded by 148 amino acids with a typical UQ_con conserved domain. The qRT-PCR technology was used to analyze the gene expression level of roots, stems, leaves and flowers in potato cultivars 'Atlantic' and 'Qingshu 9'. The results showed that the gene expression level was the highest in leaves and flowers. The expression level in 'Atlantic' was significantly higher than 'Qingshu 9'. Under drought and salt stress, potato StUBC12 gene up-regulated to increase plant stress resistance. This study could provide a theoretical basis for further study of the potato stress-resistance mechanism and the functional of the StUBC12 gene.

Since the start of the development strategy of potato staple food in China, breeding of new varieties of main grain potato has become another new goal of breeding. In this study, on the basis of morphological identification, the genetic relationship of 99 potato (Solanum tuberosum) varieties (lines) was studied by SSR marker technology. 32 pairs of SSR primers with high polymorphism index were screened and the tested materials were amplified by SSR analysis. On the basis of 12 morphological traits, 99 potato germplasms were cluster analyzed according to UPGMA (unweighted pair group method analysis) based on Euclidean distance. Morphological cluster results showed that the tested materials were divided into 2 categories at a Euclidean distance of 6.79. ClassⅠcontained 90 materials, accounting for 91 % of the tested materials; Category Ⅱ contained 9 materials including Lishu6, Qingshu2, Red rose, 1025, Dongnong310, Ganguzi, Zhongshu 21, Yunshu 606, Foreign 2. SSR analysis showed that a total of 202 isometric points were detected in 32 pairs of SSR primers, and the polymorphism ratio reached to 100 %. The genetic similarity coefficients (GS) of the tested materials were 0.627~0.926. When GS was 0.627, the tested materials were grouped into 2 categories. There were 27 genotypes in the first category, accounting for 27 % of the tested materials; 72 genotypes in the second category, accounting for 73%; in this group, the 2 varieties of Lishu6 and Huaen1 were separated separately which had larger genetic differences from other genotypes. At the GS 0.637, Kondor, Mira and Kexin22 were separated separately from the other 70 materials. The results of morphological and SSR clustering showed that the genetic difference of potato varieties in the same incubated unit was small. Analyzing the clarity of amplified bands, the number of polymorphic bands and the repeatability of primers, a total of 6 pairs of SSR core primers STI005, STI022, STI024, STI041, SSR223 and STG0016 were identified for constructing the fingerprint profiles of 99 materials. Combining these 6 pairs of core primers, 99 varieties could be distinguished completely. According to STI005 / STI024/STG0016/SSR223/STI022/ STI041, the fingerprints of 99 varieties were constructed. The results showed that the cluster analysis of morphological markers and SSR markers were consistent in the group division and genetic background, and the differences of morphological traits could reflect the differences of gene level to some extent. This study provides a reference for the identification of potato varieties and the selection of excellent hybrid combinations.

Peanut (Arachis hypogaea) was regarded to have the characteristic of moderate salt tolerance. It has become an important way during the full utilization of saline and alkali soil by searching salt-tolerant related genes, creating salt-tolerant germplasms and breeding salt-tolerant varieties in peanut. The peanut transcriptome database of salt tolerant mutant and mutagenic parent was used to screen the differentially expressed genes of late embryogenesis abundant proteins (Lea) family, and AhLea-D was obtained successfully. In order to make a further study on the role of the AhLea-D in peanut salt tolerance, the full length cDNA sequence of AhLea-D (GenBank No. MN393179) containing 534 bp was obtained by PCR amplification, which encoded an acidic protein with 177 amino acids, molecular weight 17.89 kD, theoretical isoelectric point 5.78. AhLea-D protein had average coefficient of hydrophilicity -0.983, instability index less than 40, and it belonged to the stable hydrophilic protein. AhLea-D was ligated to plasmid PCAMBIA1301 to construct the over-expression recombinant vector PCAMBIA1301-AhLea-D, which was then transferred to peanut variety 'Huayu23' by Agrobacterium-mediated pollen tube channel method. The pods from the infected flowers were harvested and the seeds were identified by PCR amplification method. The result showed the positive rate of transformation was about 52%. The expression level of AhLea-D gene was detected by qRT-PCR amplification, and the expression level of the transgenic plants were found to be 7~12 times higher than that of the non-transformed control plants. Transgenic plants and controls were treated with salt stress by irrigating 250 mmol/L NaCl. After a week of stress treatment, the control leaves showed obvious wilting and yellowing, while the transgenic plants showed no obvious changes. The photosynthesis parameter including photosynthetic net rate (Pn), stomatal conductance (Gs), intercellular CO₂ concentration (Ci), and transpiration rate (Tr) were analyzed and determined, which showed that the Pn, Gs and Tr of transgenic plants were higher than those of the non-transgenic plants, and the Ci was lower in transgenic plants than non-transgenic plants. The superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) activities and malonic dialdehyde (MDA) content of the plants were also analyzed and determined, which showed that the activities of SOD, POD and CAT of transgenic plants were significantly higher than those of control plants (P<0.05), and the content of MDA was lower than that of the control plants. Based on the above results, it was speculated that over-expression of the AhLea-D gene increased the salt tolerance of the transgenic peanut plants by maintaining Pn and photosynthetic efficiency, and by keeping the activities of antioxidant enzymes to eliminate free radicals. The results might provide theoretical basis for developing a new stress-resistant breeding approach in peanut, and also provide a new salt tolerance candidate gene.

Pectin methylesterase (PME) is a cell wall-localized protein associated with aluminum tolerance of plant. Tamba Black soybean (Glycine max cv. Tamba) belongs to leguminous plants and has great potential for aluminum tolerance. To study the function of GmPME2 (GenBank No. MN867684) in Tamba Black soybean under aluminum stress, this research cloned one of Tamba Black soybean's gene of GmPME2 by using reverse transcription PCR (RT-PCR). The coding regions of GmPME2 was 900 bp in length and encoded a protein of 299 amino acids with the isoelectric point of 8.85. GmPME2 was a stable protein with instability coefficient of 30.11<40. The expression analyses of GmPME2 in Tamba Black soybean challenged by Al³⁺ (pH4.5, 0.5 mmol/L CaCl₂, 50 μmol/L AlCl₃) showed an upregulation of GmPME2 within 12 h and downregulation from 12~24 h with the highest spot on the 12 h. Expression of GmPME2 in roots was significantly higher than that in stems or leaves (P<0.05), especially in root tips. Transient expression of tobacco (Nicotiana benthamiana) revealed that GmPME2 protein was located in cell wall. The expression vector, pBI121-GmPME2-eGFP, was constructed and transformed into tobacco by Agrobacterium mediated transformation and subsequently obtained transgenic tobaccos. Three transgenic tobaccos (GmPME2-1, GmPME2-3 and GmPME2-4) were selected to investigate the aluminum tolerance, The results showed that the relative expression of GmPME2 increased significantly (P<0.05), GmPME2 activity and malondialdehyde (MDA) content in root tips had remarkable decrease in root relative elongation compared to wild type (WT). Hematoxylin and Evans blue staining revealed a deeper stain in transgenic tobacco than that in WT. Compared with WT, the secretion of citrate in transgenic tobacco root tips significantly increased due to more aluminum absorption (P<0.05). This research indicated that plant could enhance its aluminum tolerance by decreasing aluminum absorption in root tips via regulating the expression of PME2 gene which provides the genetic resources for researches in aluminum toxicity.

ζ-carotene desaturase (ZDS) is the rate-limiting enzyme in carotenoid synthesis, catalyzing ζ-carotene to form lycopene. In this study, a ZDS gene was cloned from Hibiscus esculentus by transcriptome sequencing. The full-length cDNA sequence of ZDS was 2 189 bp, which contained a 1 686 bp ORF that encoded 561 amino acids, with a predicted molecular weight of 61.79 kD and a hypothetical isoelectric point (pI) of 6.807. It shared over 90% identity with the homologous proteins from Gossypium arboreum, G. hirsutum, Corchorus capsularis, Durio zibethinus and Theobroma cacao, showing that it was highly conservative. This gene was named HeZDS and the GenBank No. was MG372371. qRT-PCR analysis revealed that HeZDS could be expressed in different tissues of H. esculentus, including roots, stems, leaves, flowers, fruits and pod of Hibiscus esculentus, and the highest expression was found in mature leaves during leaf growth and tender pods (2 d after anthesis) in fruit development, respectively. Moreover, the expression of HeZDS gene had close correlations with carotenoid contents. These results provide a basis for the study of the HeZDS gene function and carotenoid regulation mechanism in H. esculentus.

As a phytoalexin produced by plants under biotic and abiotic stresses, resveratrol is a metabolite of stilbene synthases (STSs). Chinese wild Vitis quinquangularis accession 'Danfeng-2' contains a high concentration of resveratrol in its fruits, which also possesses strong plant resistance against diseases. The expressions and functions study of its STS genes can serve as the disease-resistant germplasm resources for resistance breeding. Based on the previous findings, this study explored the expression patterns of STS genes VqSTS12, VqSTS24 and VqSTS25 through a variety of biotic and abiotic stress treatments. As the qRT-PCR results revealed, the VqSTS12, VqSTS24 and VqSTS25 genes could be induced and expressed significantly just after 12 h of powdery mildew induction. Besides, their second-highest expressions were found during 96~120 h. Additionally, the 3 STS genes could be expressed under the following stress treatments: methyl jasmonate (MeJA), salicylic acid (SA), abscisic acid (ABA), trauma, low temperature, salt and drought. This indicated that the VqSTS12, VqSTS24 and VqSTS25 genes could be broadly induced in response to biotic and abiotic stresses. Subcellular localization in leaves of Nicotiana benthamiana observations showed that these 3 STSs were localized in the cytoplasm of leaf epidermal cells. In Vitis vinifera cv. 'Thompson Seedless' overexpressing VqSTS12 and VqSTS25, significant up-regulations of STS genes (VqSTS12 and VqSTS25) were found in transgenic plants that were under drought stress simulated with 10% PEG6000. Moreover, the physiological indices like malondialdehyde (MDA) content, peroxidase (POD) activity and catalase (CAT) activity of the transgenic plants were all superior to the wild-type plants, which suggested that the over-expression of VqSTS12 and VqSTS25 genes improved the drought resistance of Thompson Seedless. In summary, the STS genes VqSTS12, VqSTS24 and VqSTS25 could be used as the disease-resistant germplasm resources of improved European grape varieties for resistance breeding. The results of the present study provide the candidate genes and referential basis for further improving the disease and stress resistances of Vitis vinifera.

Microcystin-LR (MC-LR) is a widespread toxin produced by Cyanobacteria that pollutes the water environment. Animal studies provide strong evidence of positive associations between microcystins (MCs) exposure and reproductive toxicity, representing a threat to human reproductive health and the biodiversity of wild life. However, little is known about its toxic effects on animal oocytes. This study evaluated the toxic effects of MC-LR on porcine (Sus scrofa) oocyte in vitro maturation. The cumulus-oocyte complexes (COCs) were exposed to 0, 20, 40, 60 μg/mL MC-LR for maturation culture. Immunofluorescent staining coupled with confocal microscopy imaging techniques was used to examine the chromosomes alignment and spindle tubulin distribution of the MC-LR treated oocytes. The results showed that the maturation rate of pig oocytes exposed to MC-LR decreased in a dose-dependent manner. After 44 h in culture, most of the control oocytes had extruded the first polar body and arrested at the MⅡ stage in vitro, and the first polar body extrusion rate was 72.81%. However, the rate was significantly reduced to 56.71% (P<0.05) and 45.00% (P<0.01) when treated with 40 and 60 μg/mL of MC-LR, respectively. After MC-LR (60 μg/mL) treatment, the proportion of the oocytes arrested in telophase Ⅰ (TⅠ) stage was significantly increased from 10.63% to 45.49% (P<0.001), and the proportion of the oocytes showed obvious defects in homologous chromosome segregation and alignmen was also increased from 11.94% to 44.33% (P<0.001). Further subcellular structure examination results showed that the proportion of oocytes with spindle structure disorder increased significantly after MC-LR treatment. The results of this study indicated that MC-LR had obvious toxic damage to the spindle structure of porcine oocyte, resulting in abnormal alignment of homologous chromosomes and arrestment of meiosis in TⅠ stage, eventually resulted in failure of porcine oocyte maturation (P<0.01). The results will provide an experimental basis for further understanding the reproductive toxicity and its mechanism of MC-LR, and also provide experimental reference for effective prevention and treatment of diseases related to the reproductive toxicity of MC-LR

The regulation of angiogenic factors in the ovary of sheep (Ovis aries) is closely related to vascular proliferation, follicular development and corpus luteum formation, and has an important regulatory effect on the reproductive performance of sheep. In this study, qRT-PCR was used to detect the expression of VEGF gene in different ovarian ovary, and the expression and distribution of VEGF protein in ovarian tissue was analyzed by Immunohistochemistry and Western blot combined with IPP(image pro plus) technology. The results showed that VEGF was expressed in sheep ovary, and the expression of high fertility sheep was significantly higher than that of low fertility sheep (P<0.05). Studies showed that VEGF-positive signals were mainly localized in granulosa cells, luteal cells and vascular endothelial cells in different follicles of sheep, and were also weakly expressed in ovarian stroma and atresia follicular granulosa cells, the integrated optical density analysis of VEGF found that The expression of VEGF in the ovarian tissues of Small-tailed Han sheep, Hu sheep and Tibetan sheep was consistent. During the development of different follicles, the integrated optical density values of the follicles gradually increased and synchronized with follicular development. The positive signal strength was consistent with the follicle size, and the expression in the early stage of luteal formation was significantly higher than that in the degenerative stage. Researches found that high expression of VEGF could induce more angiogenesis around the follicles and corpus luteum in sheep, which improved the permeability of microvessels and capillaries and played an active role in increasing the number and quality of dominant follicles. This study provides basic information for revealing some of the genetic mechanisms of differences in sheep fecundity.

Early pregnancy factor (EPF), a kind of pregnancy related protein, plays an important role in the development of embryo as an autocrine and paracrine immune tolerance factor and growth factor of trophoblast layer. In this study, ovary, uterus, fallopian tube and placenta samples were collected respectively from female yak (Bos grunniens) during follicular phase, luteal phase and early pregnant period to investigate the expression of EPF and it role in reproduction. qRT-PCR, Western blott and immunohistochemistry (IHC) techniques were used to detect the expression of EFP mRNA and protein. The result indicated that EPF protein was most distributed in uterine glandular epithelial cells, endometrial epithelial cells, oviduct epithelial cells, membranous luteal cells, placental trophoblastic giant cells and crypt epithelial cells. qRT-PCR results showed that expression level of EPF mRNA in the ovary was significantly lower during follicular phase than that during early pregnant period and luteal phase. In oviduct and uterus the expression level of EPF mRNA was significantly higher during the follicular phase than that during the luteal phase and early pregnant period. The expression of EPF mRNA in ovarian and maternal placenta during early pregnant period was significantly higher than other reproductive organs. Western blot results showed that the expression of EPF protein in ovarian and oviduct during the early pregnant period was significantly higher than that during the luteal phase and follicular phase. In uterus of the follicular phase, the expression of EPF protein was significantly higher than that during the early pregnant period and luteal phase. It was noticeable that the expression of EPF in ovary and maternal placenta in early pregnant period was significantly higher than other reproductive organs, especially in the ovary. The results showed that EPF was spatiotemporal differently expressed in the main reproductive organs of female yak. During early pregnancy, EPF was mainly derived from ovary and maternal placenta. This study provides some useful materials to investigate the whole role of EPF in reproduction of yak.

Bacillus thuringiensis (Bt) Cry toxin gene has been used to cultivate insect-resistant crop varieties, which has brought great demonstration effects to the application of protein insecticidal materials. Cadherin is one of the potential receptors of Cry toxin, carrying out target screening is of great significance for the screening and creation of novel insecticidal protein materials. In this study, Humanized Single Domain Antibody Library was screened to generate single-domain antibodies against cadherin-like protein of rice leaf roller (Cnaphalocrocis medinalis) with cadherin-like protein toxin binding region (TBR) as the antigen by phage display technique. The substructive selection strategy was used to enrich phage single domain antibodies that specifically recognized TBR protein after 4 rounds of adsorption-elution-amplification. Single colonies were randomly picked from the 4th round of screening and identified by monoclonal ELISA. Four positive clones containing the complete single domain antibody gene fragment were obtained. After sequencing and amino acid sequence alignment, 3 positive clones with different sequences were finally obtained (B4, C11 and E8). The 3 positive clones identified by Western blot and ELISA binding tests had high binding activity to cadherin-like protein from rice leaf roller. Competitive inhibition test showed that 100 μg/mL Cry1Ac could inhibit 28.6% binding of C11 to the cadherin-like protein of rice leaf roller. In this study, phage display technique was used to screen phage single domain antibodies against cadherin-like protein of rice leaf roller, which could provide material basis for further development of mimics with Cry toxin functional and biological activities.

Ustilago esculenta can infect Zizania latifolia to induce edible fleshy stem, which called Jiaobai in China. It was found that U. esculenta was a dimorphic fungus, there is transformation between yeast type and mycelium type, and this dimorphic transition was related to its pathogenicity. When the lipopeptide pheromone encoded by mfa (mating factor) gene and pheromone receptor encoded by the pra (pheromone receptor) gene recognize each other, they activate cyclic adenosine monophosphate-protein kinase A (cAMP-PKA) signal transduction pathway and mitogen-activated protein kinase cascade (MAPK) signaling pathways, which make the fusion of 2 yeast-type compatible strains and form hyphae, then have pathogenicity. In this study, the gene Uegpa3 (Ustilago esculenta guanine uncleotide-binding protein subunit alpha-3)(GenBank No. ALS87611.1), encoding the α subunit of G protein, was cloned based on the whole genome analysis of U. esculenta. The cDNA length of Uegpa3 is 1 065 bp, without introns, encoding 354 amino acids, with a typical Gα domain. The expression patterns analysis showed that the expression of Uegpa3 was up-regulated during conjunction tube formation. Uegpa3 deletion mutant was constructed based on PEG (polyethylene glycol) mediated protoplast transformation and it was found that the Uegpa3 deletion strain could not fuse to form mycelium. Furthermore, the ability of conjunction tube was lost in Uegpa3 mutant during mating and the expression of pheromone synthesis gene was significantly inhibited. The above results demonstrated that Uegpa3 affected the conjugation tube formation in the dimorphic transition in U. esculenta through regulating phenome recognition. This study preliminarily explored the function of the Uegpa3, a gene encoding one of the G protein α subunits in U. esculenta, and discussed its role in dimorphic transition, which provide basic material for the research on the interaction mechanism between the U. esculenta and Z. latifolia.

Root rot is the main disease that leads to soybean (Glycine max) yield reduction. Preliminary work screened a strain named Bacillius velezensis 3A3-15 with strong antagonistic effect on Fusarium oxysporum which is pathogenic bacteria of root rot in the soybean. To investigate the effect of biocontrol B. velezensis 3A3-15 on bacterial community structure in rhizosphere soil of potted soybean, the pots containing F. oxysporum were divide into 2 groups, the experimental group used 3A3-15 agent in the pot soil and the control group did not use 3A3-15 agent. High throughput sequencing technique was used to determine the bacterial community structure of the rhizosphere soil. Taxonomy was used to annotate the OTU (operational taxonomic units) after grouping by using the usearch61 clustering method. Alpha diversity analysis showed that Shannon index, PD (phylogenetic diversity) value, Simpson index and Chao1 index in the test group decreased, but no significance. The test group had 2 258 OTUs (operational taxonomic units) specifically, decreased by 32.74% when compared with the control group (3357). Top 10 dominant microbial bacteria at phylum level were Proteobacteria, Acidobacteria, Actinobacteria, Gemmatimonadetes, Bacteroidetes, Chloroflexi, Verrucomicrobia, Planctomycetes, TM7 and Nitrospirae. There was no significant difference in the abundance of dominant phylum between the test and control group. However, the abundance of Archaea, Tracer, Fibrobacteria and OP11 in the test group was significantly higher than that in the control group (P<0.05), and the abundance of GN02 in the test group was significantly higher than that in the control group (P<0.01). The dominant species in the 2 groups were Kaistobacter and Sphingomonas. NMDS (nonmetric multidimensional scaling) analysis showed that the difference of bacterial community in test group was smaller than that in control group. In conclusion, the addition of Bacillus velezensis 3A3-15 increased the abundance of some non-dominant bacteria and reduced the differences of bacterial community structure, but these changes were no significant in the total abundance and diversity of soil bacteria between 2 groups. The above results could provide reference data for studying on biological control mechanism and safe application of 3A3-15 biocontrol agent.

With the continuous deepening of the safety management of agricultural genetically modified organisms (GMOs) in China, the role of technical standards has become more prominent, and it has become an important technical support for legal management. After 16 years of hard work, China has basically established a safety standards system for agricultural GMOs. By December 2018, the Ministry of Agriculture/Ministry of Agriculture and Rural Affairs had released 201 safety standards for agricultural GMOs, and 193 were currently in force. This paper summarizes the status quo of China's agricultural GMOs safety standards system, analyzes the characteristics of agricultural GMOs safety standards system and raises the existing problems, and discusses the development direction of agricultural GMOs safety standards system construction in the future. This paper provides reference for the further improvement of China's agricultural GMOs safety standards, which will provide better technical support for the safety management of agricultural GMOs.

Milk fat is an important evaluation index for milk quality. In addition to serving as an energy source, milk fat plays an important physiological role in human health. Mammary fatty acid metabolism in ruminants is regulated by nuclear transcription factors. In recent years, the regulation of nuclear transcription factors in ruminant mammary fatty acid metabolism has made important progress. A decisive role of ligand-dependent transcription factors including peroxisome proliferator-activated receptor family (PPAR), liver X receptor (LXR) in regulating mammary fatty acid metabolism was confirmed. Both PPAR and LXR can form transcriptional complexes that bind to the promoter of downstream target genes, thereby affecting the activity of pathways related to milk fat metabolism. The physiological functions of different gene subtypes of PPAR and LXR are varied. Sterol regulatory element-binding protein 1 (SREBP1), regulating de novo fatty acid synthesis, is a classical transcription factor regulating mammary fatty acid synthesis. In this paper, the recent advances in mammary fatty acid synthesis regulated by key transcription factors were reviewed, which will provide references for the regulation of milk fat synthesis.

Rice (Oryza sativa), one of the most important food crops, feeds more than 50% of the world's population. Mesocotyl is the first internode of rice seedlings when grown in dark. Rice varieties with longer mesocotyl will come up out of soil easier under direct seeding conditions. In previous studies, 5 quantitative trait loci (QTLs) regulating the length of mesocotyl have been identified from 'Kasalath' (O. sativa ssp. indica cv. Kasalath) with long mesocotyl, one of which has been fine mapped into gaoyao1 gene (GY1) (LOC_Os01g67430). Compared with dominant GY1^Nip from 'Nipponbare' (O. sativa ssp. japonica cv. Nipponbare), a G→T mutation occurred at 376 bp of the coding region of recessive gy1^Kasa in 'Kasalath', which resulted in the elongation of the mesocotyl. gy1^Kasa exists in a variety of natural germplasm resources, all of whom exhibits long mesocotyl phenotype. In order to promote the application of gy1^Kasa in breeding of rice suitable for direct seeding, a molecular marker was developed by Amplification Refractory Mutation System (ARMS) to identify the functional nucleotide polymorphism (FNP) of GY1. The molecular marker, denominated as gy1fnp, contained 4 primers located in the conserved flanking region of SNP376. A 470 bp fragment could be amplified by gy1fnpaf and gy1fnpar in both genotypes; a 194 bp fragment could only be amplified in homozygous gy1^Kasa (FNP site was T) by gy1fnpbf and gy1fnpar; a 315 bp fragment could only be amplified in GY1^Nip (FNP site was G) by gy1fnpaf and gy1fnpbr. All 3 fragments could be amplified in heterozygous genotype. The results of agarose gel electrophoresis showed that different genotypes of GY1 could be distinguished accurately and clearly. Sanger sequencing showed that the difference of banding patterns was caused by SNP376. Investigation of F₂ population of 'Kasalath' / 'Nipponbare' showed that homozygous gy1^Kasa identified by gy1fnp linked with long mesocotyl phenotype. Taken together, it was concluded that gy1fnp developed in this study could accurately distinguish 2 alleles and heterozygous genotypes of GY1 recognized by SNP276 using only one PCR reaction followed by agarose gel electrophoresis with convenience, rapidness and low cost. Marker gy1fnp could promote the application of gy1^Kasa in direct seeding rice breeding by molecular Marker Assisted Selection (MAS). Marker gy1fnp could also identify the genotype of GY1 in rice germplasm resources quickly, which was of great significance for genetic analysis and breeding practice. In addition, the successful development of gy1fnp was helpful to reduce genetic noise caused by GY1 and facilitated map based on cloning of other mesocotyl genes.

Optimizing a flow cytometry method technical scheme for determining genome size and ploidy of local pear (Pyrus) varieties in Xinjiang can detect and identify the genome size and ploidy more efficiently. Different source materials (dry method to store new shoot tender leaves, annual branch hydroponic tender leaves, autumn shoots tender leaves, seed cultivation cotyledon and seed cultivation tender leaves) of local pear varieties in Xinjiang, different mechanical dissociation methods (blade chopped method, grinding method), membrane filtration times (500 mesh filter once and filter twice) and propidium iodide (PI) dyeing liquid dosage (200 and 300 μL) were compared based on the detection results that taken diploid 'Dangshansuli' as reference. The results showed that the genomic size estimation and chromosome ploidy identification of 29 local pear varieties in Xinjiang were carried out by using the optimized detection method. The optimization method as follow: Young leaves which from annual branches hydroponic tender leaves in spring were collected 0.1 g, washed with distilled water and deionized water for 2 to 3 times, respectively. Then wipe it dry and put into a culture dish which had been added with 1 mL precooled woody plant buffer (WPB) dissociation solution, and chopped up quickly with a sharp blade. After chopping, add 1 mL WPB dissociation solution, mix and stand for 2~5 min. The mixture liquid was filtered with 500 mesh membrane into 2.0 mL centrifuge tube and incubated at 4 ℃ for 5 min. The single cell nuclear suspension were got from the treated mixture liquid by centrifuge at 4 ℃ at 1 000 r/min for 5 min. Retain 0.1 mL supernate at bottom and add 200 μL precooled WPB, 200 μL precooled PI, mix well. Place it in refrigerator at 4 ℃ and dye for 10 min without light. After staining, detect it with flow cytometer, and collect 5 000 mononuclear particle granules. Using this method, the results showed that the mean genome size of local pear varieties in Xinjiang detected was (566.5±125.17) Mb. The genome size of 'Yilikamut' was (666.80±154.56) Mb, and it was defined chimera for its chromosome peaks of diploid and triploid appeared simultaneously. The genome sizes of 'Kotoamut', 'Aiwenqieke', and 'Heisuanli' were (721.12±20.87) Mb, (791.33±36.84) Mb and (725.16±76.40) Mb, respectively, which were triploid. The genome size of 'Sha01' was (1032.61±49.41) Mb, which was tetraploid. Among the 24 diploid local pear varieties in Xinjiang, the genome size ranged from (480.95±16.24) to (599.14±38.36) Mb. Among the 3 triploid, the genome size ranged from (721.12±20.87) to (791.33±36.84) Mb. In this study, flow cytometry was optimized to estimate the genomic size and identify the ploidy of local pear varieties in Xinjiang. It overcame the disadvantages of the traditional dyeing method, such as difficulty and time consuming, and the defect was that more detailed chromosome characteristic information could not be obtained. The results of this study can provide technical support for ploidy breeding and genomics research of pear in the future.

Moschus chrysogaster (sifanicus) viral hemorrhagic disease (McVHD) is an acute and highly lethal viral infectious disease, and can be a serious threat to the health and safety of musk deer population in China. McVHD is caused by Moschus chrysogaster hemorrhagic disease virus (McHDV) whose genome is highly homologous with Rabbit hemorrhagic disease virus (RHDV) which belongs to Caliciviridae Lagovirus. To develop a rapid and simple diagnostic method for McVHD, a pair of specific primers McVHD-F/R were designed and synthesized based on the comparative analysis of whole genome sequence of McHDV and RHDV in GenBank. Based on the optimization of reaction condition and analysis of specificity and sensitivity, the reverse transcription PCR (RT-PCR) method for McVHD diagnosis was established. The results showed that a specific 448 bp fragment was amplified from McHDV genome using the established RT-PCR method. The RT-PCR method was highly specific and hypersensitive, and had no cross-reaction?with?other 10 pathogenic microorganisms such as Salmonella, Pasteurella multocida, Klebsiella pneumoniae. The RT-PCR was?able?to?detect?as?low?as?10^-5 ng/μL (2.38×; 10³ copies/μL) of?McHDV RNA.?The detection results?of?clinical?samples?revealed?that?the?diagnostic method was able to effectively and quickly detect viral nucleic acid in clinical samples, and the results were completely consistent with the traditional pathogen separation identification. The RT-PCR method established in this study could provide technical support for diagnosis and molecular epidemiological survey of McVHD.