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 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
[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
The doubled haploid(DH)technique accelerates homozygosity by inducing chromosome doubling in haploid embryos derived from hybrid plants.This approach offers significant advantages over conventional rice breeding metho...The doubled haploid(DH)technique accelerates homozygosity by inducing chromosome doubling in haploid embryos derived from hybrid plants.This approach offers significant advantages over conventional rice breeding methods by shortening the breeding cycle and enabling rapid development of pure homozygous lines.Anther culture(AC)has been established as an efficient and successful method for producing DH plants via androgenesis in rice.However,despite its success in japonica rice.展开更多
Primordial germ cells(PGCs)are the stem-cell population of adult animal gametes,which develop into sperm or eggs.It can be propagated in vitro and injected into the host chicken for genome editing to obtain germline c...Primordial germ cells(PGCs)are the stem-cell population of adult animal gametes,which develop into sperm or eggs.It can be propagated in vitro and injected into the host chicken for genome editing to obtain germline chimeric chicken.However,it has the limitation that the host embryo contains endogenous PGCs,which raises complications,resultantly donor PGCs fail to compete,and transmission efficiency reduced.Therefore,to increase the transmission efficiency,we generated a novel sterile chicken with the inducible elimination of endogenous PGCs in the host.This is the first study that applied the herpes simplex virus thymidine kinase(HSV-TK)cell ablation system in avian.CRISPR/Cas9-mediated homology-directed repair was performed to localize the HSV-TK suicide gene to the last exon of the deleted in azoospermialike(DAZL)gene,and ganciclovir(GCV)was added to induce the apoptosis in the germ cells of the host embryo.The sterilized host embryo introduced genome-edited PGCs to produce chimeric chicken carrying exogenous germ cells only.It was observed that the germline transmission efficiency was 100%achieved,and the obtained chicks were purely from donor breeds.The technologies established in the current study have important applications in germplasm conservation and gene editing in chicken.展开更多
The embryo rescue technique plays an essential role in developing new seedless grape varieties.To enhance the efficiency of seedless grape embryo rescue breeding,this study evaluated 22 hybrid combinations and systema...The embryo rescue technique plays an essential role in developing new seedless grape varieties.To enhance the efficiency of seedless grape embryo rescue breeding,this study evaluated 22 hybrid combinations and systematically investigated the effects of parental genotypes and plant hormones on embryo development and germination.Additionally,an in-depth analysis was conducted on the conversion of abnormal plantlets.Results indicate that‘Ruby Seedless’,‘Delight’,‘Huozhouheiyu’,‘Zitian Seedless’,and‘Zhengyan Seedless’are suitable as maternal parents,whereas‘Zitian Seedless’,‘Shennongxiangfeng’,‘Hongqitezao’,and‘Guibao’perform optimally as paternal parents.Among these,the crosses‘Ruby Seedless×Shennongxiangfeng’and‘Ruby Seedless×Zitian Seedless’exhibited the highest embryo rescue efficiency,with embryo development rates of 55.05 and 59.76%,yielding 1,348 and 2,235 viable plantlets,respectively.When 1.0 mg L^(–1) zeatin (ZT) was added to the MM3 medium supplemented with 0.2 mg L^(–1) indole-3-acetic acid (IAA),the embryo development rate of‘Ruby Seedless×Zitian Seedless’increased by 64.73%.In the WPM germination medium,supplementation with 0.2 mg L^(–1) ZT and 0.2 mg L^(–1) IAA resulted in the highest germination rate of 85.71%for the hybrid combination‘Huozhouheiyu×Shine Muscat’.Furthermore,3,365 abnormal plantlets were rescued via direct transformation and hypocotyl-induced adventitious bud regeneration,among which 1,234 were transformed into normal plantlets.Following hybridization,a total of 4,287 plants were successfully acclimatized and transplanted.This study provides theoretical insights to improve the efficiency of embryo rescue breeding in seedless grapes and offers valuable genetic resources for future breeding programs.展开更多
Background:Chronic endometritis(CE)is an important pathological factor contributing to female infertility and recurrent pregnancy loss.Although antibiotics are the primary clinical treatment for CE,they do not effecti...Background:Chronic endometritis(CE)is an important pathological factor contributing to female infertility and recurrent pregnancy loss.Although antibiotics are the primary clinical treatment for CE,they do not effectively improve pregnancy outcomes.Wen Yang Hua Zhuo(WYHZ)is a clinically employed classical formula known for its effects in warming yang,tonifying the spleen and kidneys,and resolving dampness.However,its underlying mechanisms remain unclear.This study aimed to elucidate how WYHZ modulates the immunometabolic microenvironment at the maternal-fetal interface in CE by targeting the MCT/HIF-1α/LDHA pathway to promote embryo implantation.Methods:In vivo,the model of CE was established by intrauterine injection of lipopolysaccharide(LPS)(1 mg/mL)into female C57/BL mice,followed by WYHZ treatment for 3 weeks to evaluate its effects on embryo implantation.Mechanistic studies were further conducted using the MCT-1 inhibitor AZD3965 and adeno-associated virus-mediated HIF-1αknockdown.In vitro,an in vitro CE model consisting of M1 macrophages and Ishikawa,as well as an in vitro embryo implantation model mediated by JAR cells,were constructed using Transwell,and the therapeutic mechanisms of WYHZ was validated using AZD3965 and lentiviral sh HIF-1αintervention.Metabolic enzyme activity assays,protein antibody microarrays,immunofluorescence,Western blotting,Seahorse analysis,and ELISA were employed.Results:WYHZ improved the immune-inflammatory microenvironment at the maternal-fetal interface by reducing pro-inflammatory cytokines and increasing anti-inflammatory factors.In parallel,WYHZ reprogrammed endometrial metabolism by enhancing glycolysis and suppressing mitochondrial oxidative phosphorylation,thereby improving endometrial receptivity and embryo implantation.Mechanistically,WYHZ activated the MCT/HIF-1α/LDHA pathway in endometrial epithelial cells,alleviating inflammatory stress and restoring receptivity.Both AZD3965 intervention and HIF-1αknockdown impaired endometrial receptivity and implantation,effects that were reversed by WYHZ.Conclusion:WYHZ modulates the immunometabolic microenvironment of the endometrium in the context of CE by targeting the activation of the MCT/HIF-1α/LDHA pathway,which improves endometrial receptivity and promotes embryo implantation.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
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 (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.
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 the Science and Technology Innovation Program of Fujian Agriculture and Forestry University,China(Grant No.KFB22045)the General Program of Natural Science Foundation Fujian,China(Grant No.2023J01460).
文摘The doubled haploid(DH)technique accelerates homozygosity by inducing chromosome doubling in haploid embryos derived from hybrid plants.This approach offers significant advantages over conventional rice breeding methods by shortening the breeding cycle and enabling rapid development of pure homozygous lines.Anther culture(AC)has been established as an efficient and successful method for producing DH plants via androgenesis in rice.However,despite its success in japonica rice.
基金supported by the National Key R&D Program of China(2021YFD1300100)Guangxi Key R&D Program,China(AB21220005)+1 种基金Reproductive Medicine,Guangxi Medical and Health Key Discipline Construction Project of the Affiliated Hospitalthe National Natural Science Foundation of China(32360180)。
文摘Primordial germ cells(PGCs)are the stem-cell population of adult animal gametes,which develop into sperm or eggs.It can be propagated in vitro and injected into the host chicken for genome editing to obtain germline chimeric chicken.However,it has the limitation that the host embryo contains endogenous PGCs,which raises complications,resultantly donor PGCs fail to compete,and transmission efficiency reduced.Therefore,to increase the transmission efficiency,we generated a novel sterile chicken with the inducible elimination of endogenous PGCs in the host.This is the first study that applied the herpes simplex virus thymidine kinase(HSV-TK)cell ablation system in avian.CRISPR/Cas9-mediated homology-directed repair was performed to localize the HSV-TK suicide gene to the last exon of the deleted in azoospermialike(DAZL)gene,and ganciclovir(GCV)was added to induce the apoptosis in the germ cells of the host embryo.The sterilized host embryo introduced genome-edited PGCs to produce chimeric chicken carrying exogenous germ cells only.It was observed that the germline transmission efficiency was 100%achieved,and the obtained chicks were purely from donor breeds.The technologies established in the current study have important applications in germplasm conservation and gene editing in chicken.
基金supported by the China Agriculture Research System of MOF and MARA (CARS-29-yc-3)。
文摘The embryo rescue technique plays an essential role in developing new seedless grape varieties.To enhance the efficiency of seedless grape embryo rescue breeding,this study evaluated 22 hybrid combinations and systematically investigated the effects of parental genotypes and plant hormones on embryo development and germination.Additionally,an in-depth analysis was conducted on the conversion of abnormal plantlets.Results indicate that‘Ruby Seedless’,‘Delight’,‘Huozhouheiyu’,‘Zitian Seedless’,and‘Zhengyan Seedless’are suitable as maternal parents,whereas‘Zitian Seedless’,‘Shennongxiangfeng’,‘Hongqitezao’,and‘Guibao’perform optimally as paternal parents.Among these,the crosses‘Ruby Seedless×Shennongxiangfeng’and‘Ruby Seedless×Zitian Seedless’exhibited the highest embryo rescue efficiency,with embryo development rates of 55.05 and 59.76%,yielding 1,348 and 2,235 viable plantlets,respectively.When 1.0 mg L^(–1) zeatin (ZT) was added to the MM3 medium supplemented with 0.2 mg L^(–1) indole-3-acetic acid (IAA),the embryo development rate of‘Ruby Seedless×Zitian Seedless’increased by 64.73%.In the WPM germination medium,supplementation with 0.2 mg L^(–1) ZT and 0.2 mg L^(–1) IAA resulted in the highest germination rate of 85.71%for the hybrid combination‘Huozhouheiyu×Shine Muscat’.Furthermore,3,365 abnormal plantlets were rescued via direct transformation and hypocotyl-induced adventitious bud regeneration,among which 1,234 were transformed into normal plantlets.Following hybridization,a total of 4,287 plants were successfully acclimatized and transplanted.This study provides theoretical insights to improve the efficiency of embryo rescue breeding in seedless grapes and offers valuable genetic resources for future breeding programs.
基金supported by the National Natural Science Foundation of China(grant number:82205172,82274570).
文摘Background:Chronic endometritis(CE)is an important pathological factor contributing to female infertility and recurrent pregnancy loss.Although antibiotics are the primary clinical treatment for CE,they do not effectively improve pregnancy outcomes.Wen Yang Hua Zhuo(WYHZ)is a clinically employed classical formula known for its effects in warming yang,tonifying the spleen and kidneys,and resolving dampness.However,its underlying mechanisms remain unclear.This study aimed to elucidate how WYHZ modulates the immunometabolic microenvironment at the maternal-fetal interface in CE by targeting the MCT/HIF-1α/LDHA pathway to promote embryo implantation.Methods:In vivo,the model of CE was established by intrauterine injection of lipopolysaccharide(LPS)(1 mg/mL)into female C57/BL mice,followed by WYHZ treatment for 3 weeks to evaluate its effects on embryo implantation.Mechanistic studies were further conducted using the MCT-1 inhibitor AZD3965 and adeno-associated virus-mediated HIF-1αknockdown.In vitro,an in vitro CE model consisting of M1 macrophages and Ishikawa,as well as an in vitro embryo implantation model mediated by JAR cells,were constructed using Transwell,and the therapeutic mechanisms of WYHZ was validated using AZD3965 and lentiviral sh HIF-1αintervention.Metabolic enzyme activity assays,protein antibody microarrays,immunofluorescence,Western blotting,Seahorse analysis,and ELISA were employed.Results:WYHZ improved the immune-inflammatory microenvironment at the maternal-fetal interface by reducing pro-inflammatory cytokines and increasing anti-inflammatory factors.In parallel,WYHZ reprogrammed endometrial metabolism by enhancing glycolysis and suppressing mitochondrial oxidative phosphorylation,thereby improving endometrial receptivity and embryo implantation.Mechanistically,WYHZ activated the MCT/HIF-1α/LDHA pathway in endometrial epithelial cells,alleviating inflammatory stress and restoring receptivity.Both AZD3965 intervention and HIF-1αknockdown impaired endometrial receptivity and implantation,effects that were reversed by WYHZ.Conclusion:WYHZ modulates the immunometabolic microenvironment of the endometrium in the context of CE by targeting the activation of the MCT/HIF-1α/LDHA pathway,which improves endometrial receptivity and promotes embryo implantation.
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.
