Bacterial infections of avian embryos can lead to an increase in embryo mortality,and the proliferation of antimicrobial-resistant bacteria aggravates the situation.A low hatching rate also poses a challenge to the po...Bacterial infections of avian embryos can lead to an increase in embryo mortality,and the proliferation of antimicrobial-resistant bacteria aggravates the situation.A low hatching rate also poses a challenge to the population of artificially bred Crested Ibises(Nipponia nippon).This study aims to determine the potential association between bacterial infection and the death of Crested Ibis embryos,and whether there is convergence between antimicrobial resistance and virulence in strain.In this study,13 Escherichia coli and 12 Proteus mirabilis isolates were recovered from dead Crested Ibis embryos.The pathogenicity examination confirmed the pathogenicity of all isolates,and multiple virulence genes detected by PCR-sequencing demonstrated the presence of irp2 and iuc D(100%),fim C and iss(92.31%)in E.coli,and uca A(58.33%)in P.mirabilis.Antimicrobial susceptibility test demonstrated that isolates were mainly resistant to amoxicillin(E.coli:76.92%,P.mirabilis:91.67%),cefazolin(E.coli:76.92%,P.mirabilis:91.67%),oxytetracycline(E.coli:92.31%,P.mirabilis:75.00%)and sulfamethoxazole-trimethoprim(E.coli:53.85%,P.mirabilis:33.33%),and more than 30%of isolates showed multidrug-resistance(MDR).Further analyses detected extended-spectrumβ-lactamase(ESBL)genes,of which blaTEM-1(E.coli:100%,P.mirabilis:100%)had the highest frequency,followed by the blaCTX-M-55(E.coli:92.31%,P.mirabilis:50%),blaCTX-M-14(E.coli:76.92%,P.mirabilis:33.33%),blaCTX-M-65(E.coli:15.38%,P.mirabilis:16.67%),and all isolates were negative for blaSHV and blaOXA.Pearson's correlation analysis showed a positive correlation between the presence ofβ-lactam resistance and ESBL genes,while mainly negative correlations were observed between the presence of ESBL genes and virulence genes.Furthermore,the conjugation experiment and PFGE revealed that the isolates were primarily polyclonal,and there was horizontal transfer of resistance or virulence genes by plasmids.Based on the results,E.coli and P.mirabilis were responsible for embryonic mortality of the ibises in this study.The co-presence and co-transfer of ESBL genes and virulence genes can pose a potential threat to the health of the Crested Ibis,and measures such as prudent use of antimicrobials,and constant surveillance of resistance and pathogenicity,must be implemented at the Crested Ibis breeding base.展开更多
Litchi has great economic significance as a global fruit crop.However,the advancement of litchi functional genomics has encountered substantial obstacles due to its recalcitrance to stable transformation.Here,we prese...Litchi has great economic significance as a global fruit crop.However,the advancement of litchi functional genomics has encountered substantial obstacles due to its recalcitrance to stable transformation.Here,we present an efficacious Agrobacterium tumefaciens-mediated transformation system in somatic embryos of‘Heiye'litchi.This system was developed through the optimization of key variables encompassing explant selection,A.tumefaciens strain delineation,bacterium concentration,infection duration,and infection methodology.The subsequent validation of the transformation technique in litchi was realized through the ectopic expression of LcMYB1,resulting in the generation of transgenic calli.However,the differentiation of transgenic calli into somatic embryos encountered substantial challenges.To delineate the intricate molecular underpinnings of LcMYB1's inhibitory role in somatic embryo induction,a comprehensive transcriptome analysis was conducted that encompassed embryogenic calli(C),globular embryos(G),and transgenic calli(TC).A total of 1,166 common differentially expressed genes(DEGs)were identified between C-vs.-G and C-vs.-TC.Gene Ontology(GO)annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis revealed that these common DEGs were mostly related to plant hormone signal transduction pathways.Furthermore,RT-qPCR corroborated the pronounced down-regulation of numerous genes that are associated with somatic embryo induction within the transgenic calli.The development of this transformation system provides valuable support for functional genomics research in litchi.展开更多
[Objective]This study was to screen out suitable genotypes and basic medium for the culture of maize mature embryos.[Method]Using mature embryos of nine maize genotypes as explants,the effects of genotypes and basic m...[Objective]This study was to screen out suitable genotypes and basic medium for the culture of maize mature embryos.[Method]Using mature embryos of nine maize genotypes as explants,the effects of genotypes and basic medium on callus induction and subculture were investigated.[Result]The genotypes performed better in callus induction and subculture were found in turn 853-35,853-209,Dan 34 and 81162.MS medium is better than N6 medium in the callus induction from maize embryos,while N6 medium is more suitable for callus subculture.[Conclusion]Our study further improved the tissue culture system in maize with mature embryos as explants.展开更多
In this study we studied the factors influencing the callus induction from mature embryos of maize inbred lines Qi 319, Zhen 58, Chang 7 -2, Lx 9801 and 81162, such as genotype, combination of plant growth regulators,...In this study we studied the factors influencing the callus induction from mature embryos of maize inbred lines Qi 319, Zhen 58, Chang 7 -2, Lx 9801 and 81162, such as genotype, combination of plant growth regulators, and low-temperature pretreatment. The results showed that the induction rate of Qi 319 was the highest among the four genotypes tested; combination of 4.0 mg/L 2,4-D ＋ 0.5 mg/L 6-BA was suitable for inducing callus from mature embryos; three days of 4℃ pretreatment can promote the callus induction significantly. The indices optimized in the present study are helpful for establishing genetic transformation system in maize without considering seasonal variation.展开更多
Fluctuations of levels of several endogenous plant hormones in isolated rice ( Oryza sativa ssp. japonica) embryos during early and mid-embryogenesis and early stages of germination were studied by enzyme-linked immun...Fluctuations of levels of several endogenous plant hormones in isolated rice ( Oryza sativa ssp. japonica) embryos during early and mid-embryogenesis and early stages of germination were studied by enzyme-linked immunosorbent assay (ELISA). Embryos were collected at different days after pollination (DAP) and different days after imbibition (DAI) of mature seeds. The contents of gibberellin(1) (GA(1)), abscisic acid (ABA), isopentenyladenine and isopentenyladenosine ( iPAs), zeatin and zeatin riboside ( ZRs) were immunochemically assayed. The GA(1) level was the highest among all hormones tested. The variations of GA(1) levels were opposite with the ABA levels, with some exceptions. During early and mid-embryogenesis, the levels of GA(1) and ABA were the highest at 4 DAP. From 8 to 18 DAP, GA(1) level declined, whereas the ABA level increased. During germination, GA(1) level increased at 2 DAI whereas simultaneously the ABA content decreased. The highest ratio of GA(1)/ABA was observed at 2 DAI The levels of iPAs and ZRs were maxima in the embryos at 4 DAP, decreased to a very low level and maintained constant thereafter. Our results provide further evidence that GA(1) plays an important role in the early stages of embryo development and germination. The balance between GA(1) and ABA, rather than their absolute contents, controls these processes throughout the development, whereas iPAs and ZRs may play important roles in early embryogenesis. The use of isolated embryos as starting material avoids the usual interferences with other tissues such as the endosperm. In addition, this is the first report dealing with the hormonal balance of early-embryos in rice.展开更多
An in situ hybridization technique for localization of calmodulin(CaM) mRNA in isolated entire embryo sacs and proembryos in Nicotiana tabacum L.cv.W38 has been developed. This technique can be applied to smal...An in situ hybridization technique for localization of calmodulin(CaM) mRNA in isolated entire embryo sacs and proembryos in Nicotiana tabacum L.cv.W38 has been developed. This technique can be applied to small amounts of materials in which a whole view of CaM mRNA distribution can be obtained. The authors revealed that CaM mRNA expression changes dramatically before and after fertilization. Especially interesting is that a prominent CaM mRNA band appears between the egg apparatus and polar nuclei temporarily during the period of pollination and fertilization. The band disappears just prior to fertilization and expands to a fan_shaped region that occupies the micropylar portion of the embryo sac. After fertilization, CaM mRNA accumulates in the elongated zygotes with higher concentration in their chalazal portion than in the micropylar portion. Such an asymmetrical pattern continues to manifest in the early proembryos. It is supposed that CaM mRNA may be involved in the early events and signaling steps associated with double fertilization and zygote polarization in higher plants.展开更多
In ecological zone of Chengdu, Sichuan, microspore culture was carried out in Brassica napus L. to study the influencing factors on microspore culture. The results showed that the temperature on microspore formation s...In ecological zone of Chengdu, Sichuan, microspore culture was carried out in Brassica napus L. to study the influencing factors on microspore culture. The results showed that the temperature on microspore formation stage, day and night temperature, disinfection solution of buds, cultivation concentration on microspore and strain-age were both important influencing factors on microspore culture. At a temperature below 5 ℃ or above 20 ℃, the material had a much lower embryo producing rate or even could not produce any embryo, but at the optimum temperature of 10 -15 ℃ the embryo yield was up to 300 pieces per bud; the best cultivation effect appeared when 0. 1% HgCl2 was used for disin- fection; the best density of microspore was 3 -4 buds per dish; In 2009, while strain-age was from 125 d to 150 d, the microspore embryo yield increased as strain-age increased. When stain-age was 150 days, the microspore embryo yield was up to the highest, but the yield declined after 150 days.展开更多
[Objective]The aim was to study the effective factors on culturing Prunus salicina cv.Zaoshi embryos in vitro.[Method]Different age,culture medium and GA3 which affected culturing of prumssalicina embryos were discuss...[Objective]The aim was to study the effective factors on culturing Prunus salicina cv.Zaoshi embryos in vitro.[Method]Different age,culture medium and GA3 which affected culturing of prumssalicina embryos were discussed.[Result]The result showed that on the part of 26-41 d,after the blooming period,the embryos remained tiny and retained endosperms and showed no signs of change after having cultured for three generations.On the part of 48 d,after the blooming period,the endosperms had disappeared,the embryos kept growing until they filled the seed cavity;when they were planted on the MS culture medium,their survival rate reached 77%,in its first generation,the response of embryos was discernible.On the part of 65 d,after the blooming period,4.5% of their embryos grew into shoots on the MS culture medium;with the age of embryos growing,the survival rate of shoots increased until it reached 26% when the fruits went into ripeness;the embryos produced calli in their first generation of culturing.On the part of 65-83 d,after the blooming period,the embryos produced calli through more than 2-3 generations.On the part of 88 d,after the blooming period,the survival rate of shoots on the WPM culturing base doubled compared with that on the MS culturing base;on the same culture medium,the embryos were inhibited from growing into shoots when BA,KT or 2,4-D was added on to the culture medium.The survival rate of shoots was increased remarkably when the seeds were treated in 1 000 mg/L GA3.[Conclusion] This study provided experimental basis for the establishment of Prunus salicina cv.Zaoshi,embryos rescue techniques and cross breading.展开更多
[Objective] The aim was to explore the conditions of high frequency induction of embryonic callus and plant regeneration of maize. [Method] Immature embryos of maize inbred lines were used as explants to study the eff...[Objective] The aim was to explore the conditions of high frequency induction of embryonic callus and plant regeneration of maize. [Method] Immature embryos of maize inbred lines were used as explants to study the effect of different 2,4-D concentrations on the induction of callus,the effect of different 6-BA concentrations on the differentiation of test-tube plantlet,as well as the effect of different IBA concentrations on the rooting of test-tube plantlet. [Result] 2,4-D showed obvious effect on the induction of inducement rate of maize,and the optimum induction medium was：N6 ＋ 2 mg/L of 2,4-D ＋ 500 mg/L of CH ＋ 500 mg/L of Pro ＋10 mg/L of AgNO3; the optimum differentiation medium was：N6 ＋ 0.5 mg/L of BA ＋ 500 mg/L of Pro; the optimum for the rooting of test-tube plantlet was 1/2 MS ＋ 0.5 mg/L of IBA. [Conclusion] The study had provided basis for the genetic transformation of maize.展开更多
[Objective] The aim of this study was to manually isolate the microstructures such as ovule and embryo of Capsella bursa-pastoris (L.) Medic. by the non-enzymatic method. [Method] With the gum block and dissecting mic...[Objective] The aim of this study was to manually isolate the microstructures such as ovule and embryo of Capsella bursa-pastoris (L.) Medic. by the non-enzymatic method. [Method] With the gum block and dissecting microscope, its floral characteristics, ovule and embryo were manually isolated and observed by the non-enzymatic method. [Result] C. bursa-pastoris had four calyces, four petals, tetradynamous stamen and silicle, while its embryogenesis contained globular embryo, heart-shaped embryo and torpedo-shaped embryo. [Conclusion] The microstructures such as ovule and embryo of plants become easier to be isolated manually with the viscosity of gum block, which has small damage on embryo or disadvantages of keeping original shapes, and also can used in multi fields studies. This test can also be accomplished in labs with poor experiment facilities.展开更多
In culturing early mouse embryos in vitro,liquid paraffin and alcohol exert deleterious influence on the development of embryos. Some of light liquid paraffin produced by Chinese factories have proved harmful for earl...In culturing early mouse embryos in vitro,liquid paraffin and alcohol exert deleterious influence on the development of embryos. Some of light liquid paraffin produced by Chinese factories have proved harmful for early mouse embryos. As shown by our experiments, the nitronaphthalene contained and the specific gravity of liquid paraffin were not involved in the injurious effects.However,alcohol mingled in medium had harmful effects on the development of embryos. At the 0.1% concentration of alcohol in medium the proportion of embryos developing to blastocysts decreased to 73.9%. When the concentration of alcohol was increased to 0.8%, all embryos ceased developing. In our experiments, CO_2 which contained 0.13% alcohol had no visible effects on the development of embrvos in vitro.展开更多
Green fluorescent protein ( GFP ) gene was expressed transiently in 2-3 d old rice embryos by electroporation with the aid of a specially designed loading net. Under suitable conditions (500 μF capacitance, 300 V/c...Green fluorescent protein ( GFP ) gene was expressed transiently in 2-3 d old rice embryos by electroporation with the aid of a specially designed loading net. Under suitable conditions (500 μF capacitance, 300 V/cm Voltage, 100 μg/mL plasmid DNA), the percentage of embryos expressing GFP was up to 35%. The highest electroporation efficiency (40%) was obtained at pH 5.8 of the electroporation buffer. The GFP gene driven by the Ubi promoter produced the highest efficiency. Thus, on the basis of optimizing electroporation conditions, a transformation system has been developed for young embryos in rice. The electroporated 4-6 d old embryos regenerated plantlets under the controlled cultural conditions. Fluorescence microscopic observations indicated that GFP gene expressed in their calli and R0 plantlets.展开更多
In previous study we reported that pretreatment with plasmolysis enhanced somatic embryo formation in hypocotyls of Eleutherococcus senticosus.In the present study,the expression level of callose synthase gene in embr...In previous study we reported that pretreatment with plasmolysis enhanced somatic embryo formation in hypocotyls of Eleutherococcus senticosus.In the present study,the expression level of callose synthase gene in embryos of E.senticosus in response to 2,4-D,sucrose and mannitol treatments was analyzed by RT-PCR.The results show that plasmolysis pretreatment using sucrose and mannitol significantly promoted the expression of callose synthase gene.Also,the thicker cell walls of explant plasmolyzed compared with controls were observed during the somatic embryogenesis.We suggest that the callose may make the cells in epidermis separate from neighboring cells and then develop into embryogenic potential cells.展开更多
The cattle different stage embryos obtained from in vitro was studied using the technology of single preimplantation embryo mRNA different display:single 8-cell and blastocyst stage embryos were studied using technolo...The cattle different stage embryos obtained from in vitro was studied using the technology of single preimplantation embryo mRNA different display:single 8-cell and blastocyst stage embryos were studied using technology of mRNA different display and one different fragment was found. The result suggested that this fragment displayed high homology (99%) to cattle mRNA for ribosomal protein L31. Then to detect the expression of RPL31mRNA in 8 cell and blastocyst stage embryos by real-time quantitative PCR,the result showed the relative amount of 8 cells was 3.2 times of blastocyst's.展开更多
In recent years,with the wide applications and mineral exploitation of rare earth elements,their potential environmental and health effects have caused increasing public concern.Effect of rare earth elements La and Yb...In recent years,with the wide applications and mineral exploitation of rare earth elements,their potential environmental and health effects have caused increasing public concern.Effect of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos were studied.The embryos were exposed to La3＋ or Yb3＋ at 0,0.01,0.1,0.3,0.5 and 1.0 mmol/L,respectively.Early life stage parameters such as egg and embryo mortality,gastrula development,tail detachment,eyes,somite formation,circulatory system,pigmentation,malformations,hatching rate,length of larvae and mortality were investigated.The results showed La3＋ and Yb3＋ delayed zebrafish embryo and larval development,decreased survival and hatching rates,and caused tail malformation in a concentration-dependent way.Moreover,heavy rare-earth ytterbium led to more severe acute toxicity of zebrafish embryo than light rare-earth lanthanum.展开更多
Wheat, one of the most important food crops, has been extensively studied with respects to plant regeneration and transformation employing the immature embryos as recipient tissues. However, the transformed tissues of...Wheat, one of the most important food crops, has been extensively studied with respects to plant regeneration and transformation employing the immature embryos as recipient tissues. However, the transformed tissues often become severely necrotic after co-cultivation with Agrobacterium, which is one of the major obstacles in gene delivery. In this study, wheat varieties CB037, Kenong 199, Xinchun 9, Lunxuan 987, and Shi 4185 showed desirable culture potential or high infection ability in Agrobacterium-mediated transformation. Similarly, optimal regeneration conditions were determined by testing their ability to inhibit the cell necrosis and cell death phenotype. Two auxins of 2,4-dichlorophenoxyacetic acid （2,4-D） and 3,6-dichloro-o-anisic acid （dicamba） were evaluated for highly significant effect on both callus and plantlet production, although they were genotype-dependent in wheat. Substitution of 2,4-D by dicamba enhanced the growth and regeneration ability of callus from the immature embryos of most genotypes tested. The callus growth state couldn’t be modified by adding antioxidants such as ascorbic acid, cysteine, and silver nitrate or organic additives such as thiamine HCl and asparagine to the media. On the contrary, the best tissue statement and plant regeneration was achieved by employing the media containing the simplest MS （Murashige and Skoog） components and dicamba without organic components and vitamins. Thereby, our results are thought to inhibit wheat cell necrosis effectively and suggested to be used for more wheat genotypes.展开更多
The immature embryos （IEs） of wheat are the most widely used tissues for in vitro culture and genetic transformation due to its high regeneration competency. However, this explant can only be maintained in 4℃ daily...The immature embryos （IEs） of wheat are the most widely used tissues for in vitro culture and genetic transformation due to its high regeneration competency. However, this explant can only be maintained in 4℃ daily cooler for a short period time for its use in plant tissue culture or transformation experiments. This study aimed to investigate the effects of environmental temperature, cryopreservation storage temperature, and heat shock culture （HSC） temperature on the regeneration frequency of wheat IEs. Results indicated that environmental temperature significantly affected the induction of embryonic calli. The optimum total accumulated temperature （TAT） during the time of anthesis and sampling for regeneration of these tissues was around 280℃ for spring wheat type cv. CB037 and approximately 300℃ for winter wheat type cv. Kenong 199. Regeneration ability obviously declined when the highest environmental temperature was over 35℃ for 1 d or a high temperature between 30 and 33℃ lasted for 5 d during anthesis and sampling. This finding was verified by culturing the freshly isolated IEs under different temperatures from 29 to 37℃ in different controlled growth incubators for 5 d; the IEs almost completely lost regeneration ability when the temperature rose to 37℃. Cryopreservation of-20℃ caused the wheat samples lost ability of producing callus or embryonic callus in a few days, and cryopreservation of-10℃ more than 10 d made the regeneration potential of the tissues dramatically declined. Comparatively, the temperature that best maintained high regeneration ability was -5℃, at which the materials can be maintained for around 1 mon. In addition, the preservation of the immature samples at -5 or -10℃ inhibited the direct germination of the IEs, avoiding the embryo axis removing process. Our results are useful for ensuring that field collection and cryopreservation of the wheat IEs are done correctly to enable tissue culture and genetic transformation.展开更多
Oocyte quality has long been considered as a main limiting factor for in vitro fertilization （IVF）. In the past decade, extensive observations demonstrated that the mitochondrion plays a vital role in the oocyte cyt...Oocyte quality has long been considered as a main limiting factor for in vitro fertilization （IVF）. In the past decade, extensive observations demonstrated that the mitochondrion plays a vital role in the oocyte cytoplasm, for it can provide adenosine triphosphate （ATP） for fertilization and preimplantation embryo development and also act as stores of intracellular calcium and proapoptotic factors. During the oocyte maturation, mitochondria are characterized by distinct changes of their distribution pattern from being homogeneous to heterogeneous, which is correlated with the cumulus apoptosis. Oocyte quality decreases with the increasing maternal age. Recent studies have shown that low quality oocytes have some age-related dysfunctions, which include the decrease in mitochondrial membrane potential, increase of mitochondrial DNA （mtDNA） damages, chromosomal aneuploidies, the incidence of apoptosis, and changes in mitochoncLrial gene expression. All these dysfunctions may cause a high level of de- velopmental retardation and arrest of preimplantation embryos. It has been suggested that these mitochondrial changes may arise from excessive reactive oxygen species （ROS） that is closely associated with the oxidative energy production or calcium overload, which may trigger permeability transition pore opening and subsequent apoptosis. Therefore, mitochondria can be seen as signs for oocyte quality evaluation, and it is possible that the oocyte quality can be improved by enhancing the physical function of mitochondria. Here we reviewed recent advances in mitochondrial functions on oocytes.展开更多
As the most widely used plasticizers in the world, phthalate esters （PAEs） are potential endocrine disruption compounds （EDCs）. In the present study, the toxicity of dimethyl phthalate （DMP）, diethyl phthalate ...As the most widely used plasticizers in the world, phthalate esters （PAEs） are potential endocrine disruption compounds （EDCs）. In the present study, the toxicity of dimethyl phthalate （DMP）, diethyl phthalate （DEP）, dibutyl phthalate （DBP）, di （2-ethylhexyl） phthalate （DEHP） on embryogenesis and larvae development of the marine univalve Haliotis diversicolor supertexta was examined in laboratory. The results show that the malformation of embryos appeared during the experiment, such as embryos died or lysed, small transparent flocculent rings studded on the periphery of the embryo, and the larvae could failed to hatch. In embryo toxic test, embryos incubated at the highest concentration of DMR DEP and DBP solutions showed significantly high abnormal rate compared with the control, while DEHP solutions displayed no significant difference. In larval toxic test, in all concentrations of DMP, DEP and DBP solutions, larval settlement rates were low significantly than that of the control. Similarly, DEHP solutions show nearly no effect on the larval settlement. The order of toxicity on embryos and larvae is DBP〉DEP〉DMP〉DEHE Being a simple and easy stimulation to indoor spawn, sensitive to environmental factors, and short culture time, the embryos of H. diversicolor supertexta can be used to indicate toxicity of the PAEs.展开更多
Objective This study was aimed to investigate the toxic effects of 3 nanomaterials, i.e. multi-walled carbon nanotubes （MWCNTs）, graphene oxide （GO）, and reduced graphene oxide （RGO）, on zebrafish embryos. Metho...Objective This study was aimed to investigate the toxic effects of 3 nanomaterials, i.e. multi-walled carbon nanotubes （MWCNTs）, graphene oxide （GO）, and reduced graphene oxide （RGO）, on zebrafish embryos. Methods The 2-h post-fertilization （hpf） zebrafish embryos were exposed to MWCNTs, GO, and RGO at different concentrations （1, 5, 10, 50, 100 mg/L） for 96 h. Afterwards, the effects of the 3 nanomateria on spontaneous movement, heart rate, hatching rate, length of larvae, mortality, and malformations Is were evaluated. Results Statistical analysis indicated that RGO significantly inhibited the hatching of zebrafish embryos. Furthermore, RGO and MWCNTs decreased the length of the hatched larvae at 96 hpf. No obvious morphological malformation or mortality was observed in the zebrafish embryos after exposure to the three nanomaterials. Conclusion MWCNTs, GO, and RGO were all toxic to zebrafish embryos to influence embryos hatching and larvae length. Although no obvious morphological malformation and mortality were observed in exposed zebrafish embryos, further studies on the toxicity of the three nanomaterials are still needed.展开更多
基金supported by Research on Breeding and Healthy Breeding Technology of Xueyu White Chicken(mating line)in Tibet Science and Technology Program(XZ202101ZY0002N)the National Key R&D Program Project(2022YFD1600902-4)Sichuan Province Regional Innovation Cooperation Project(2023YFQ0050)。
文摘Bacterial infections of avian embryos can lead to an increase in embryo mortality,and the proliferation of antimicrobial-resistant bacteria aggravates the situation.A low hatching rate also poses a challenge to the population of artificially bred Crested Ibises(Nipponia nippon).This study aims to determine the potential association between bacterial infection and the death of Crested Ibis embryos,and whether there is convergence between antimicrobial resistance and virulence in strain.In this study,13 Escherichia coli and 12 Proteus mirabilis isolates were recovered from dead Crested Ibis embryos.The pathogenicity examination confirmed the pathogenicity of all isolates,and multiple virulence genes detected by PCR-sequencing demonstrated the presence of irp2 and iuc D(100%),fim C and iss(92.31%)in E.coli,and uca A(58.33%)in P.mirabilis.Antimicrobial susceptibility test demonstrated that isolates were mainly resistant to amoxicillin(E.coli:76.92%,P.mirabilis:91.67%),cefazolin(E.coli:76.92%,P.mirabilis:91.67%),oxytetracycline(E.coli:92.31%,P.mirabilis:75.00%)and sulfamethoxazole-trimethoprim(E.coli:53.85%,P.mirabilis:33.33%),and more than 30%of isolates showed multidrug-resistance(MDR).Further analyses detected extended-spectrumβ-lactamase(ESBL)genes,of which blaTEM-1(E.coli:100%,P.mirabilis:100%)had the highest frequency,followed by the blaCTX-M-55(E.coli:92.31%,P.mirabilis:50%),blaCTX-M-14(E.coli:76.92%,P.mirabilis:33.33%),blaCTX-M-65(E.coli:15.38%,P.mirabilis:16.67%),and all isolates were negative for blaSHV and blaOXA.Pearson's correlation analysis showed a positive correlation between the presence ofβ-lactam resistance and ESBL genes,while mainly negative correlations were observed between the presence of ESBL genes and virulence genes.Furthermore,the conjugation experiment and PFGE revealed that the isolates were primarily polyclonal,and there was horizontal transfer of resistance or virulence genes by plasmids.Based on the results,E.coli and P.mirabilis were responsible for embryonic mortality of the ibises in this study.The co-presence and co-transfer of ESBL genes and virulence genes can pose a potential threat to the health of the Crested Ibis,and measures such as prudent use of antimicrobials,and constant surveillance of resistance and pathogenicity,must be implemented at the Crested Ibis breeding base.
基金supported by the National Natural Science Foundation of China(31872066 and 32272663)the Science and Technology Planning Project of Guangzhou,China(2023B01J2002)+2 种基金the Key Research and Development Program of Hainan,China(ZDYF2023XDNY052)the Seed Industry Engineering Project of Department of Agriculture and Rural Affairs of Guangdong,China(2022-NPY-00-004 and 2022-NBH00-001)the Litchi Industry Science and Technology Special Mission of Yunnan,China(202204BI090021)。
文摘Litchi has great economic significance as a global fruit crop.However,the advancement of litchi functional genomics has encountered substantial obstacles due to its recalcitrance to stable transformation.Here,we present an efficacious Agrobacterium tumefaciens-mediated transformation system in somatic embryos of‘Heiye'litchi.This system was developed through the optimization of key variables encompassing explant selection,A.tumefaciens strain delineation,bacterium concentration,infection duration,and infection methodology.The subsequent validation of the transformation technique in litchi was realized through the ectopic expression of LcMYB1,resulting in the generation of transgenic calli.However,the differentiation of transgenic calli into somatic embryos encountered substantial challenges.To delineate the intricate molecular underpinnings of LcMYB1's inhibitory role in somatic embryo induction,a comprehensive transcriptome analysis was conducted that encompassed embryogenic calli(C),globular embryos(G),and transgenic calli(TC).A total of 1,166 common differentially expressed genes(DEGs)were identified between C-vs.-G and C-vs.-TC.Gene Ontology(GO)annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis revealed that these common DEGs were mostly related to plant hormone signal transduction pathways.Furthermore,RT-qPCR corroborated the pronounced down-regulation of numerous genes that are associated with somatic embryo induction within the transgenic calli.The development of this transformation system provides valuable support for functional genomics research in litchi.
基金Supported by National Natural Science Foundation of China(31070224)National Natural Science Foundation of China(30970219)Key Project from Science and Technology Department in Jilin Province(20080203)~~
文摘[Objective]This study was to screen out suitable genotypes and basic medium for the culture of maize mature embryos.[Method]Using mature embryos of nine maize genotypes as explants,the effects of genotypes and basic medium on callus induction and subculture were investigated.[Result]The genotypes performed better in callus induction and subculture were found in turn 853-35,853-209,Dan 34 and 81162.MS medium is better than N6 medium in the callus induction from maize embryos,while N6 medium is more suitable for callus subculture.[Conclusion]Our study further improved the tissue culture system in maize with mature embryos as explants.
基金Supported by Heilongjiang August First Land Reclamation University(Establishment of fast and high-frequency regeneration system of maize)~~
文摘In this study we studied the factors influencing the callus induction from mature embryos of maize inbred lines Qi 319, Zhen 58, Chang 7 -2, Lx 9801 and 81162, such as genotype, combination of plant growth regulators, and low-temperature pretreatment. The results showed that the induction rate of Qi 319 was the highest among the four genotypes tested; combination of 4.0 mg/L 2,4-D ＋ 0.5 mg/L 6-BA was suitable for inducing callus from mature embryos; three days of 4℃ pretreatment can promote the callus induction significantly. The indices optimized in the present study are helpful for establishing genetic transformation system in maize without considering seasonal variation.
文摘Fluctuations of levels of several endogenous plant hormones in isolated rice ( Oryza sativa ssp. japonica) embryos during early and mid-embryogenesis and early stages of germination were studied by enzyme-linked immunosorbent assay (ELISA). Embryos were collected at different days after pollination (DAP) and different days after imbibition (DAI) of mature seeds. The contents of gibberellin(1) (GA(1)), abscisic acid (ABA), isopentenyladenine and isopentenyladenosine ( iPAs), zeatin and zeatin riboside ( ZRs) were immunochemically assayed. The GA(1) level was the highest among all hormones tested. The variations of GA(1) levels were opposite with the ABA levels, with some exceptions. During early and mid-embryogenesis, the levels of GA(1) and ABA were the highest at 4 DAP. From 8 to 18 DAP, GA(1) level declined, whereas the ABA level increased. During germination, GA(1) level increased at 2 DAI whereas simultaneously the ABA content decreased. The highest ratio of GA(1)/ABA was observed at 2 DAI The levels of iPAs and ZRs were maxima in the embryos at 4 DAP, decreased to a very low level and maintained constant thereafter. Our results provide further evidence that GA(1) plays an important role in the early stages of embryo development and germination. The balance between GA(1) and ABA, rather than their absolute contents, controls these processes throughout the development, whereas iPAs and ZRs may play important roles in early embryogenesis. The use of isolated embryos as starting material avoids the usual interferences with other tissues such as the endosperm. In addition, this is the first report dealing with the hormonal balance of early-embryos in rice.
文摘An in situ hybridization technique for localization of calmodulin(CaM) mRNA in isolated entire embryo sacs and proembryos in Nicotiana tabacum L.cv.W38 has been developed. This technique can be applied to small amounts of materials in which a whole view of CaM mRNA distribution can be obtained. The authors revealed that CaM mRNA expression changes dramatically before and after fertilization. Especially interesting is that a prominent CaM mRNA band appears between the egg apparatus and polar nuclei temporarily during the period of pollination and fertilization. The band disappears just prior to fertilization and expands to a fan_shaped region that occupies the micropylar portion of the embryo sac. After fertilization, CaM mRNA accumulates in the elongated zygotes with higher concentration in their chalazal portion than in the micropylar portion. Such an asymmetrical pattern continues to manifest in the early proembryos. It is supposed that CaM mRNA may be involved in the early events and signaling steps associated with double fertilization and zygote polarization in higher plants.
文摘In ecological zone of Chengdu, Sichuan, microspore culture was carried out in Brassica napus L. to study the influencing factors on microspore culture. The results showed that the temperature on microspore formation stage, day and night temperature, disinfection solution of buds, cultivation concentration on microspore and strain-age were both important influencing factors on microspore culture. At a temperature below 5 ℃ or above 20 ℃, the material had a much lower embryo producing rate or even could not produce any embryo, but at the optimum temperature of 10 -15 ℃ the embryo yield was up to 300 pieces per bud; the best cultivation effect appeared when 0. 1% HgCl2 was used for disin- fection; the best density of microspore was 3 -4 buds per dish; In 2009, while strain-age was from 125 d to 150 d, the microspore embryo yield increased as strain-age increased. When stain-age was 150 days, the microspore embryo yield was up to the highest, but the yield declined after 150 days.
基金Support by Major Science and Technology Projects of Guangdong Province (2004A20102002)~~
文摘[Objective]The aim was to study the effective factors on culturing Prunus salicina cv.Zaoshi embryos in vitro.[Method]Different age,culture medium and GA3 which affected culturing of prumssalicina embryos were discussed.[Result]The result showed that on the part of 26-41 d,after the blooming period,the embryos remained tiny and retained endosperms and showed no signs of change after having cultured for three generations.On the part of 48 d,after the blooming period,the endosperms had disappeared,the embryos kept growing until they filled the seed cavity;when they were planted on the MS culture medium,their survival rate reached 77%,in its first generation,the response of embryos was discernible.On the part of 65 d,after the blooming period,4.5% of their embryos grew into shoots on the MS culture medium;with the age of embryos growing,the survival rate of shoots increased until it reached 26% when the fruits went into ripeness;the embryos produced calli in their first generation of culturing.On the part of 65-83 d,after the blooming period,the embryos produced calli through more than 2-3 generations.On the part of 88 d,after the blooming period,the survival rate of shoots on the WPM culturing base doubled compared with that on the MS culturing base;on the same culture medium,the embryos were inhibited from growing into shoots when BA,KT or 2,4-D was added on to the culture medium.The survival rate of shoots was increased remarkably when the seeds were treated in 1 000 mg/L GA3.[Conclusion] This study provided experimental basis for the establishment of Prunus salicina cv.Zaoshi,embryos rescue techniques and cross breading.
基金Supported by Natural Science Foundation of Guangxi Province(Guikezi 0991096)~~
文摘[Objective] The aim was to explore the conditions of high frequency induction of embryonic callus and plant regeneration of maize. [Method] Immature embryos of maize inbred lines were used as explants to study the effect of different 2,4-D concentrations on the induction of callus,the effect of different 6-BA concentrations on the differentiation of test-tube plantlet,as well as the effect of different IBA concentrations on the rooting of test-tube plantlet. [Result] 2,4-D showed obvious effect on the induction of inducement rate of maize,and the optimum induction medium was：N6 ＋ 2 mg/L of 2,4-D ＋ 500 mg/L of CH ＋ 500 mg/L of Pro ＋10 mg/L of AgNO3; the optimum differentiation medium was：N6 ＋ 0.5 mg/L of BA ＋ 500 mg/L of Pro; the optimum for the rooting of test-tube plantlet was 1/2 MS ＋ 0.5 mg/L of IBA. [Conclusion] The study had provided basis for the genetic transformation of maize.
基金Supported by Natural Science Foundation (No. 30770124)SRTP Fund from Henan University of Science and Technology~~
文摘[Objective] The aim of this study was to manually isolate the microstructures such as ovule and embryo of Capsella bursa-pastoris (L.) Medic. by the non-enzymatic method. [Method] With the gum block and dissecting microscope, its floral characteristics, ovule and embryo were manually isolated and observed by the non-enzymatic method. [Result] C. bursa-pastoris had four calyces, four petals, tetradynamous stamen and silicle, while its embryogenesis contained globular embryo, heart-shaped embryo and torpedo-shaped embryo. [Conclusion] The microstructures such as ovule and embryo of plants become easier to be isolated manually with the viscosity of gum block, which has small damage on embryo or disadvantages of keeping original shapes, and also can used in multi fields studies. This test can also be accomplished in labs with poor experiment facilities.
文摘In culturing early mouse embryos in vitro,liquid paraffin and alcohol exert deleterious influence on the development of embryos. Some of light liquid paraffin produced by Chinese factories have proved harmful for early mouse embryos. As shown by our experiments, the nitronaphthalene contained and the specific gravity of liquid paraffin were not involved in the injurious effects.However,alcohol mingled in medium had harmful effects on the development of embryos. At the 0.1% concentration of alcohol in medium the proportion of embryos developing to blastocysts decreased to 73.9%. When the concentration of alcohol was increased to 0.8%, all embryos ceased developing. In our experiments, CO_2 which contained 0.13% alcohol had no visible effects on the development of embrvos in vitro.
文摘Green fluorescent protein ( GFP ) gene was expressed transiently in 2-3 d old rice embryos by electroporation with the aid of a specially designed loading net. Under suitable conditions (500 μF capacitance, 300 V/cm Voltage, 100 μg/mL plasmid DNA), the percentage of embryos expressing GFP was up to 35%. The highest electroporation efficiency (40%) was obtained at pH 5.8 of the electroporation buffer. The GFP gene driven by the Ubi promoter produced the highest efficiency. Thus, on the basis of optimizing electroporation conditions, a transformation system has been developed for young embryos in rice. The electroporated 4-6 d old embryos regenerated plantlets under the controlled cultural conditions. Fluorescence microscopic observations indicated that GFP gene expressed in their calli and R0 plantlets.
基金supported by the 948 Program of State Forestry Administration (No2009-4-26)co-sponsored by SRF for ROCS and the National Natural Science Foundation (No 30972390) of China
文摘In previous study we reported that pretreatment with plasmolysis enhanced somatic embryo formation in hypocotyls of Eleutherococcus senticosus.In the present study,the expression level of callose synthase gene in embryos of E.senticosus in response to 2,4-D,sucrose and mannitol treatments was analyzed by RT-PCR.The results show that plasmolysis pretreatment using sucrose and mannitol significantly promoted the expression of callose synthase gene.Also,the thicker cell walls of explant plasmolyzed compared with controls were observed during the somatic embryogenesis.We suggest that the callose may make the cells in epidermis separate from neighboring cells and then develop into embryogenic potential cells.
基金Supported by National "863" Project (2008AA101007)~~
文摘The cattle different stage embryos obtained from in vitro was studied using the technology of single preimplantation embryo mRNA different display:single 8-cell and blastocyst stage embryos were studied using technology of mRNA different display and one different fragment was found. The result suggested that this fragment displayed high homology (99%) to cattle mRNA for ribosomal protein L31. Then to detect the expression of RPL31mRNA in 8 cell and blastocyst stage embryos by real-time quantitative PCR,the result showed the relative amount of 8 cells was 3.2 times of blastocyst's.
基金supported by the National Natural Science Foundation of China (No. 10875136,10905062,11005118)the Ministry of Science and Technology of China (No. 2011CB933400)
文摘In recent years,with the wide applications and mineral exploitation of rare earth elements,their potential environmental and health effects have caused increasing public concern.Effect of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos were studied.The embryos were exposed to La3＋ or Yb3＋ at 0,0.01,0.1,0.3,0.5 and 1.0 mmol/L,respectively.Early life stage parameters such as egg and embryo mortality,gastrula development,tail detachment,eyes,somite formation,circulatory system,pigmentation,malformations,hatching rate,length of larvae and mortality were investigated.The results showed La3＋ and Yb3＋ delayed zebrafish embryo and larval development,decreased survival and hatching rates,and caused tail malformation in a concentration-dependent way.Moreover,heavy rare-earth ytterbium led to more severe acute toxicity of zebrafish embryo than light rare-earth lanthanum.
基金supported by the National Natural Science Foundation of China (30971776)the National Transgenic Specialized Research Program of China (2008ZX08010-004)
文摘Wheat, one of the most important food crops, has been extensively studied with respects to plant regeneration and transformation employing the immature embryos as recipient tissues. However, the transformed tissues often become severely necrotic after co-cultivation with Agrobacterium, which is one of the major obstacles in gene delivery. In this study, wheat varieties CB037, Kenong 199, Xinchun 9, Lunxuan 987, and Shi 4185 showed desirable culture potential or high infection ability in Agrobacterium-mediated transformation. Similarly, optimal regeneration conditions were determined by testing their ability to inhibit the cell necrosis and cell death phenotype. Two auxins of 2,4-dichlorophenoxyacetic acid （2,4-D） and 3,6-dichloro-o-anisic acid （dicamba） were evaluated for highly significant effect on both callus and plantlet production, although they were genotype-dependent in wheat. Substitution of 2,4-D by dicamba enhanced the growth and regeneration ability of callus from the immature embryos of most genotypes tested. The callus growth state couldn’t be modified by adding antioxidants such as ascorbic acid, cysteine, and silver nitrate or organic additives such as thiamine HCl and asparagine to the media. On the contrary, the best tissue statement and plant regeneration was achieved by employing the media containing the simplest MS （Murashige and Skoog） components and dicamba without organic components and vitamins. Thereby, our results are thought to inhibit wheat cell necrosis effectively and suggested to be used for more wheat genotypes.
基金the National Natural Science Foundation of China(30971776)the Transgenic Major Projects,Ministry of Agriculture of China(2011ZX08010-004)
文摘The immature embryos （IEs） of wheat are the most widely used tissues for in vitro culture and genetic transformation due to its high regeneration competency. However, this explant can only be maintained in 4℃ daily cooler for a short period time for its use in plant tissue culture or transformation experiments. This study aimed to investigate the effects of environmental temperature, cryopreservation storage temperature, and heat shock culture （HSC） temperature on the regeneration frequency of wheat IEs. Results indicated that environmental temperature significantly affected the induction of embryonic calli. The optimum total accumulated temperature （TAT） during the time of anthesis and sampling for regeneration of these tissues was around 280℃ for spring wheat type cv. CB037 and approximately 300℃ for winter wheat type cv. Kenong 199. Regeneration ability obviously declined when the highest environmental temperature was over 35℃ for 1 d or a high temperature between 30 and 33℃ lasted for 5 d during anthesis and sampling. This finding was verified by culturing the freshly isolated IEs under different temperatures from 29 to 37℃ in different controlled growth incubators for 5 d; the IEs almost completely lost regeneration ability when the temperature rose to 37℃. Cryopreservation of-20℃ caused the wheat samples lost ability of producing callus or embryonic callus in a few days, and cryopreservation of-10℃ more than 10 d made the regeneration potential of the tissues dramatically declined. Comparatively, the temperature that best maintained high regeneration ability was -5℃, at which the materials can be maintained for around 1 mon. In addition, the preservation of the immature samples at -5 or -10℃ inhibited the direct germination of the IEs, avoiding the embryo axis removing process. Our results are useful for ensuring that field collection and cryopreservation of the wheat IEs are done correctly to enable tissue culture and genetic transformation.
基金supported by the National Natural Science Foundation of China (No. 30772345)the Research Program of the Science and Technology Bureau of Zhejiang Province (No. 2006C33016)+1 种基金the Natural Science Foundation of Zhejiang Province (No. Y204202)the Chinese Medicine Research Program of Zhejiang Province (No. 2007CA071), China
文摘Oocyte quality has long been considered as a main limiting factor for in vitro fertilization （IVF）. In the past decade, extensive observations demonstrated that the mitochondrion plays a vital role in the oocyte cytoplasm, for it can provide adenosine triphosphate （ATP） for fertilization and preimplantation embryo development and also act as stores of intracellular calcium and proapoptotic factors. During the oocyte maturation, mitochondria are characterized by distinct changes of their distribution pattern from being homogeneous to heterogeneous, which is correlated with the cumulus apoptosis. Oocyte quality decreases with the increasing maternal age. Recent studies have shown that low quality oocytes have some age-related dysfunctions, which include the decrease in mitochondrial membrane potential, increase of mitochondrial DNA （mtDNA） damages, chromosomal aneuploidies, the incidence of apoptosis, and changes in mitochoncLrial gene expression. All these dysfunctions may cause a high level of de- velopmental retardation and arrest of preimplantation embryos. It has been suggested that these mitochondrial changes may arise from excessive reactive oxygen species （ROS） that is closely associated with the oxidative energy production or calcium overload, which may trigger permeability transition pore opening and subsequent apoptosis. Therefore, mitochondria can be seen as signs for oocyte quality evaluation, and it is possible that the oocyte quality can be improved by enhancing the physical function of mitochondria. Here we reviewed recent advances in mitochondrial functions on oocytes.
基金Supported by National Natural Science Foundation of China (No. 40476057)
文摘As the most widely used plasticizers in the world, phthalate esters （PAEs） are potential endocrine disruption compounds （EDCs）. In the present study, the toxicity of dimethyl phthalate （DMP）, diethyl phthalate （DEP）, dibutyl phthalate （DBP）, di （2-ethylhexyl） phthalate （DEHP） on embryogenesis and larvae development of the marine univalve Haliotis diversicolor supertexta was examined in laboratory. The results show that the malformation of embryos appeared during the experiment, such as embryos died or lysed, small transparent flocculent rings studded on the periphery of the embryo, and the larvae could failed to hatch. In embryo toxic test, embryos incubated at the highest concentration of DMR DEP and DBP solutions showed significantly high abnormal rate compared with the control, while DEHP solutions displayed no significant difference. In larval toxic test, in all concentrations of DMP, DEP and DBP solutions, larval settlement rates were low significantly than that of the control. Similarly, DEHP solutions show nearly no effect on the larval settlement. The order of toxicity on embryos and larvae is DBP〉DEP〉DMP〉DEHE Being a simple and easy stimulation to indoor spawn, sensitive to environmental factors, and short culture time, the embryos of H. diversicolor supertexta can be used to indicate toxicity of the PAEs.
基金supported by the Specialized Research Fund for the Doctoral Program of Higher Education(200800191013)the Fundamental Research Funds for the Central Universities
文摘Objective This study was aimed to investigate the toxic effects of 3 nanomaterials, i.e. multi-walled carbon nanotubes （MWCNTs）, graphene oxide （GO）, and reduced graphene oxide （RGO）, on zebrafish embryos. Methods The 2-h post-fertilization （hpf） zebrafish embryos were exposed to MWCNTs, GO, and RGO at different concentrations （1, 5, 10, 50, 100 mg/L） for 96 h. Afterwards, the effects of the 3 nanomateria on spontaneous movement, heart rate, hatching rate, length of larvae, mortality, and malformations Is were evaluated. Results Statistical analysis indicated that RGO significantly inhibited the hatching of zebrafish embryos. Furthermore, RGO and MWCNTs decreased the length of the hatched larvae at 96 hpf. No obvious morphological malformation or mortality was observed in the zebrafish embryos after exposure to the three nanomaterials. Conclusion MWCNTs, GO, and RGO were all toxic to zebrafish embryos to influence embryos hatching and larvae length. Although no obvious morphological malformation and mortality were observed in exposed zebrafish embryos, further studies on the toxicity of the three nanomaterials are still needed.
