A microfabricated electrical impedance spectroscopy (EIS) chip with microelectrodes was developed.The substrate and the electrodes of the chip were made of glass and gold,respectively.The experimental results demonstr...A microfabricated electrical impedance spectroscopy (EIS) chip with microelectrodes was developed.The substrate and the electrodes of the chip were made of glass and gold,respectively.The experimental results demonstrated that the EIS-chip could distinguish different solutions (physiological saline,culture medium,living cell suspension etc.) by scanning from 10Hz to 45kHz.A 6-element circuit model was used for fitting the real part and the imaginary part admittance curves of the living cell suspension.An actual circuit was also built and tested to verify the 6-element circuit model proposed.The micro-EIS chip has several advantages including the use of small sample volumes,high resolution and ease of operation.It shows good application prospects in the areas of cellular electrophysioiogy,drug screening and bio-sensors etc.展开更多
Glaucoma,characterized by a degenerative loss of retinal ganglion cells,is the second leading cause of blindness worldwide.There is currently no cure for vision loss in glaucoma because retinal ganglion cells do not r...Glaucoma,characterized by a degenerative loss of retinal ganglion cells,is the second leading cause of blindness worldwide.There is currently no cure for vision loss in glaucoma because retinal ganglion cells do not regenerate and are not replaced after injury.Human stem cell-derived retinal ganglion cell transplant is a potential therapeutic strategy for retinal ganglion cell degenerative diseases.In this review,we first discuss a 2D protocol for retinal ganglion cell differentiation from human stem cell culture,including a rapid protocol that can generate retinal ganglion cells in less than two weeks and focus on their transplantation outcomes.Next,we discuss using 3D retinal organoids for retinal ganglion cell transplantation,comparing cell suspensions and clusters.This review provides insight into current knowledge on human stem cell-derived retinal ganglion cell differentiation and transplantation,with an impact on the field of regenerative medicine and especially retinal ganglion cell degenerative diseases such as glaucoma and other optic neuropathies.展开更多
The effect of La, Ce was firstly tested on growth of the Taxus(T.) cuspidata cell suspensions, biosynthesis and release of taxol. The results indicate that the growth pattern of T.cuspidata cells is altered si...The effect of La, Ce was firstly tested on growth of the Taxus(T.) cuspidata cell suspensions, biosynthesis and release of taxol. The results indicate that the growth pattern of T.cuspidata cells is altered significantly by adding high concentration of rare earth in the medium. The lag and exponential phases of cell growth are shortened, the stationary phase disappears and the biomass fluctuates periodically during the decline phase. The rare earth compounds added in the exponential phase obviously increase the taxol biosyntheis and release yields of T.cuspidata cells, and the supplement of carbon source in the medium containing rare earth is also favorable to taxol biosynthesis.展开更多
Transplantation of activated transgenic Schwann cells or a fetal spinal cord cell suspension has been widely used to treat spinal cord injury. However, little is known regarding the effects of co-transplantation. In t...Transplantation of activated transgenic Schwann cells or a fetal spinal cord cell suspension has been widely used to treat spinal cord injury. However, little is known regarding the effects of co-transplantation. In the present study, autologous Schwann cells in combination with a fetal spinal cord cell suspension were transplanted into adult Wistar rats with spinal cord injury, and newly generated axonal connections were observed ultrastructurally. Transmission electron microscopic observations showed that the neuroblast first presented cytoplasmic processes, followed by pre- and postsynaptic membranes with low electron density forming a dense projection. The number and types of synaptic vesicles were increased. Synaptic connections developed from single cell body-dendritic synapses into multiple cell body-dendritic and dendrite-dendritic synapses. In addition, the cell organs of the transplanted neuroblast, oligodendroblast and astroblast matured gradually. The blood-brain barrier appeared subsequently. Moreover, neurofilament, histamine, calcitonin-gene-related peptides, and glial fibrillary acidic protein positive fibers were observed in the transplant region. These findings demonstrate that fetal spinal cord cells in the presence of autologous activated Schwann cells can develop into mature synapses in the cavity of injured spinal cords, suggesting the possibility of information exchange through the reconstructed synapse between fetal spinal cord cells and the host.展开更多
Deacetoxy-13-oxo sinenxan A (1) was converted to 9a-hydroxy-13-oxo-2a, 5a, 10b-triacetoxy-4(20),11-taxadiene (2) and 10b-hydroxy-13-oxo-2a,5a,9a-triacetoxy- 4(20), 11- taxadiene (3) by Ginkgo cell suspension cultures ...Deacetoxy-13-oxo sinenxan A (1) was converted to 9a-hydroxy-13-oxo-2a, 5a, 10b-triacetoxy-4(20),11-taxadiene (2) and 10b-hydroxy-13-oxo-2a,5a,9a-triacetoxy- 4(20), 11- taxadiene (3) by Ginkgo cell suspension cultures in 45% and 15% yields, respectively.展开更多
Leaves of fine Populus tomentosa genotype TC152 were used as explants to establish cell suspension lines. The effects of plant growth regulators on callus induction and establishment of cell suspension lines were stud...Leaves of fine Populus tomentosa genotype TC152 were used as explants to establish cell suspension lines. The effects of plant growth regulators on callus induction and establishment of cell suspension lines were studied. The callus induction rate was the highest on a MS solid medium supplemented with 1.0 mg·L^-1 2,4-D. A cell suspension line could be obtained by inoculating calli which were not subcultured into a MS liquid medium supplemented with 1.5 mg·L^-1 2,4-D. The best subculture medium was MS + 0.8 mg'L-1 2,4-D + 30 g·L^-1 sucrose with a subculture cycle of seven days.展开更多
A cell suspension culture of Panax ginseng which may be continuously subcultured has been established. Embryogenic callus derived from clutured young leaves was used to initiate the culture.Plant growth regulators, ba...A cell suspension culture of Panax ginseng which may be continuously subcultured has been established. Embryogenic callus derived from clutured young leaves was used to initiate the culture.Plant growth regulators, basal medium formula and carbohydrate levels were examined to determine their various effects on suspension culture cell growth and development. The best selection of plant growth regulator, hasal medium and carbohydrate level is 2 mg / L 2,4-D: 0.5 mg / L KT,MS and 3% sucrose respectively.展开更多
The standard detection hallmarks of apoptosis of Taxus cuspidata cells in suspension culture with Ce 4+ were studied. The condensation and margination of chromatin were observed under the electron microscopy....The standard detection hallmarks of apoptosis of Taxus cuspidata cells in suspension culture with Ce 4+ were studied. The condensation and margination of chromatin were observed under the electron microscopy. DNA fragmentation ranged 'DNA ladder' on agarose gel electrophoresis. TdT mediated dUTP nick end labeling (TUNEL) analysis of the cells reveals that the nuclear DNA strand breaks can be identified by labeling free 3′ OH termini. These results suggest that Ce 4+ can induce apoptosis of Taxus cuspidata cells and also indicate that there is a certain relationship between apoptosis and secondary metabolite product Taxol.展开更多
Suspension cultures initiated from callus derived from petiole explants of aspen hybrid (Populus tremuloides×P. tremula) produced somatic embryos. Callus was induced on a MS medium supplemented with 5mg·L^...Suspension cultures initiated from callus derived from petiole explants of aspen hybrid (Populus tremuloides×P. tremula) produced somatic embryos. Callus was induced on a MS medium supplemented with 5mg·L^-1 2,4-D and 0.05mg·L^-1 zeatin under light conditions. Embryogenic calli were obtained when a subsequent subculture of calli was suspended in the same basal medium with 10mg·L^-1 2,4-D. The highest number of globular embryos were induced from embryogenic calli by cell suspension culture in a MS liquid medium supplemented with 10mg·L^-1 2,4-D. Genotype and 2,4-D concentration were vital to the induction of embryogenic calli producing competent cells. Embryogenic calli for each genotype were heterogeneous. Green calli with gel-like consistency could yield more competent cells than light yellow embryogenic calli. However, some globular embryos broke into slices and some developed abnormally after one month of culture under the same or other hormonal conditions.展开更多
Populus spp.have long been used as model woody plant species for molecular biology research.However,tissues of poplar are often recalcitrant to experimental procedures for molecular studies.We generated a hormone auto...Populus spp.have long been used as model woody plant species for molecular biology research.However,tissues of poplar are often recalcitrant to experimental procedures for molecular studies.We generated a hormone autotrophic poplar suspension cell line from a hybrid of Populus alba×P.berolinensis‘Yinzhong’,named Qu-2.Qu-2 cells are suitable as a model biological system for studying woody plants.Qu-2 cells have many advantages over suspension cell lines derived so far from any other woody plants.Qu-2 cells are very easy to cultivate and can grow on several common plant culture media without the addition of any plant hormone.They show exceptionally high growth rates,reaching an approximately 150-fold increase in biomass after one week of culturing.Another important unique characteristic of Qu-2 cells is that they can be cryopreserved and readily reactivated.Qu-2 cells are suitable for molecular manipulations such as protoplast production,transient transformation,and RNA-seq analysis.Therefore,Qu-2 cells have the great potential to be an excellent model cell line in tree molecular biological research,ranging from physiology to gene function.The Qu-2 cells will be made available to the plant community for research.展开更多
Friable callus was induced when immature seeds of floribunda rose were inoculated on MS medium supplemented with 2,4-D 3.0 mg.L^-1. When transfered onto subculture media, friable callus developed into embryogenic call...Friable callus was induced when immature seeds of floribunda rose were inoculated on MS medium supplemented with 2,4-D 3.0 mg.L^-1. When transfered onto subculture media, friable callus developed into embryogenic callus, which was used to establish cell suspension lines. Cell suspensions had to be subcultured at a interval of 4-5 days at the first several culture cycles. The best subculturing cycle for the stable cell suspensions was 8-10 days. The best inoculum quantity was 1 mL PCV(Packed Cell Volume) per 40 mL culture fluid.展开更多
The activity of alcohol dehydrogenase (ADH) in cultured cells of various tobacco was determined. It was found that significant differences existed in cells of different varieties cultured under normal conditions and a...The activity of alcohol dehydrogenase (ADH) in cultured cells of various tobacco was determined. It was found that significant differences existed in cells of different varieties cultured under normal conditions and as well after treated with exogenous ethanol. The ADH activity had positive relation with the ability of the cells to catabolize exogenous ethanol, indicating that the main function of the ADH in tobacco cells was in the direction of converting ethanol to acetaldehyde.展开更多
In this work, we present an ultrastructural and physiological description of a novel chlorophyll-deficient, yellow cell line of the grass Bouteloua gracilis that develops etioplast-like plastids in presence of light (...In this work, we present an ultrastructural and physiological description of a novel chlorophyll-deficient, yellow cell line of the grass Bouteloua gracilis that develops etioplast-like plastids in presence of light (YELP). These mutant cells were compared to the parental, wild-type, highly chlorophyllous cells from which they were isolated. Growth analysis, based on fresh and dry weights, indicated that YELP accumulates biomass at a slower rate than the parental, green cells. Besides, YELP developed very low levels of photosynthetic pigments, reaching only 9.3% and 38.4% of chlorophyll a and chlorophyll b, respectively, developed by the wild-type cells. Likewise, the accessory pigments, carotenes and xanthophylls, were only synthesized at 8.0% and 5.4%, respectively, of the levels reached by the green cells. Electron microscopy revealed remarkable differences in plastid ultrastructure between the wild-type and mutant cells. Plastids of YELP were heterogeneous and smaller than those found in wild-type cells. YELP plastids were abnormal with poorly developed membrane systems that prevented the accumulation of chlorophyll and accessory pigments in the mutant cell line. We expect this novel, mutant cell line will provide new tools for studying plastid development and differentiation.展开更多
We used the leaf blade of rice (cultivars were Nonghu 6, Sugeng 2, Huyou 2 and Hanfeng) as initial material for protoplast culture, and a great number of regenerated plants were obtained. Rice seeds were sterilized an...We used the leaf blade of rice (cultivars were Nonghu 6, Sugeng 2, Huyou 2 and Hanfeng) as initial material for protoplast culture, and a great number of regenerated plants were obtained. Rice seeds were sterilized and germinated. The immature leaves were cut into 3-5 mm pieces when the third or forth leaf appeared. Leaf pieces were inoculated on MS medium with 2,4-D 4 mg/1, NAA 2mg/1 and IAA Img/1. After 2 wk culture, calli were induced and subcultured once or twice for multiplication. 3-5 g calli were transferred to the modified MS liquid medium with 2,4-D 2 mg/1 and KT 0.5mg/1 for suspension culture. Embryogenic cell suspension was established after 2 mo culture. The effect of the growth period of suspension cells on the展开更多
In vitro tissue culture of hard woody, endangered, medicinal plant Coscinium fenestratum is most challenging to plant tissue culturists. In the present study, petiole and leaf explants of Coscinium fenestratum were in...In vitro tissue culture of hard woody, endangered, medicinal plant Coscinium fenestratum is most challenging to plant tissue culturists. In the present study, petiole and leaf explants of Coscinium fenestratum were induced to form callus when cultured on vermicompost extract media along with coelomic fluid. Suspension medium was developed using vermicompost extract and coelomic fluid in 3:1 ratio. Phytochemical analysis of the alkaloid berberine was confirmed from callus, suspension cell culture and suspension medium by Thin Layer Chromatography and High Performance Liquid Chromatography. Vermicompost and its extracts with coelomic fluid have shown maximum (100 per cent) response of callus induction. Callus mass enlarged with increasing concentration of coelomic fluid and callus growth was assessed from the biomass. Incubation of culture tubes in dark supported callus development significantly. The Rf value of 0.36 confirmed the presence of berberine by Thin Layer Chromatography. Qualitative analysis confirmed the presence of alkaloid berberine with the retention time of 2.8 minutes similar to that of standard reference sample from Sigma chemicals, USA. The suspension medium turned deep yellow because of the release of the alkaloid. Vermicompost and its extracts along with coelomic fluid have shown the economical approach for micropropagation of economically and medicinally important plants.展开更多
Ricinus communis L. is a new copper hyperaccumulator growing on Tonglushan copper mine in Hubei Province, China. This study aimed to establish a suspension cell line of R. communis L. with stable and rapid growth for ...Ricinus communis L. is a new copper hyperaccumulator growing on Tonglushan copper mine in Hubei Province, China. This study aimed to establish a suspension cell line of R. communis L. with stable and rapid growth for further screening of heavy metal-resistant R. communis L. cells and breeding of hyperaccu- mulators. In this study, cell suspension culture conditions were optimized by using orthogonal experimental design with previously induced R. communis L. embyre- genic calluses as experimental materials, to establish the suspension cell line of R. communis L. Under the optimal conditions, growth curves of suspension cells and changes in pH of culture liquid were determined. The results showed that the optimal culture conditions for R. conmmnis L. suspension cell line were : MS + O. 5 rag/I, 6-BA + O. 2 mg/L NAA + 50 mg/L sucrose + 350 mg/L casein hydrolysate as basic medium, with callus inoculation amount of 2.5 g/50 ml, dark culture at (26 ±2) ℃ with shaking at 110 r/min. Under these conditions, increment of fresh weight and dry weight ofR. commun/s L. suspension cells reached the maximum of 4.56 g/(50 ml 14 d) and 0.49 g/(50 ml 16 d), respectively. Growth curves of R. communis L. suspension cells were basically in "S" shape. Each culture cycle lasted 16 d, and the rapid growth stage was from the 6th d to the 14th d. In a culture cycle, pH of the culture liquid declined first and then increased to the maximum and stabilized gradually.展开更多
3´,5´-Cyclic adenosine monophosphate(cAMP)is an important metabolite that is specifically enriched in jujube.However,the effect of cAMP on jujube cellular responses has not been comprehensively studied.Here,...3´,5´-Cyclic adenosine monophosphate(cAMP)is an important metabolite that is specifically enriched in jujube.However,the effect of cAMP on jujube cellular responses has not been comprehensively studied.Here,we established jujube cell suspension cultures and investigated the calcium influx in response to cAMP treatment through protoplast isolation and fluorescence intensity.Firstly,cAMP treatment could promote jujube growth and increase the content of endogenous cAMP.Using transcriptome analysis with transgenic Arabidopsis plants overexpressing adenylate cyclase(ZjAC)as a positive control,we identified 60 calcium-related differential expressed genes(DEGs)that contributed to the calcium signaling and inter-or intra-cellular responses.Pharmacological treatments such as cAMP and the calcium ionophore A23187 could induce ZjAC expression,the accumulation of cAMP and calcium influx in jujube cells,while ethylene glycol tetraacetic acid(EGTA)or bithionol treatment inhibited these changes.Moreover,the calcium channels and transporters in calcium influx,such as the ZjCNGC2 channel and the mitogen activated protein(MAP)kinase pathway,could be activated by cAMP treatment.In summary,our findings demonstrated that cAMP biosynthesis is dependent on calcium influx and the amplifying effect between calcium and cAMP may be involved in intracellular signal induction,which might contribute to the growth and development of jujube.展开更多
The authors used suspension cells of Populus euphratica to isolate protoplast in the present study. Protoplasts were successfully obtained after 4 hours incubation in enzyme solution containing 1 0% cellulase “o...The authors used suspension cells of Populus euphratica to isolate protoplast in the present study. Protoplasts were successfully obtained after 4 hours incubation in enzyme solution containing 1 0% cellulase “onozuka” R\|10, 0\^01% pectolyase Y\|23,0\^15% macerozyme R\|10 and 0\^1% hemicellulase at 25℃. Outward and inward single channels in plasma membrane were observed using cell\|attached recording of patch\|clamp technique. In this study, single channel records showed that more than one species of channel were obtained. These attempts in protoplast isolation and ion channel recording offers the opportunity to characterize cellular mechanisms of salt tolerance in tree species.展开更多
DNA methylation is widespread in plants and associated with plant development and defense mechanisms.However,the relationship between DNA methylation and plant secondary metabolism has rarely been reported.Here,when b...DNA methylation is widespread in plants and associated with plant development and defense mechanisms.However,the relationship between DNA methylation and plant secondary metabolism has rarely been reported.Here,when birch suspension cells were treated with 5-azacytidine(5-azaC),which blocks DNA methylation,triterpenoid accumulation was significantly promoted and antioxidant and defense enzymatic activity changed.For studying triterpenoid accumulation,0.1 mM azaC was optimal.A qRT-PCR assay revealed increased expression of genes encoding key triterpenoid biosynthetic enzymes.Evaluation of methylation polymorphisms at CCGG sites showed that the methylation level was lower in cells treated with 5-azaC.These results demonstrated that 5-azaC treatment led to an increase in the production of triterpenoids in cell cultures through a mechanism that involved in DNA methylation,which resulted in the induction of genes encoding the key enzymes.The study provides evidence of a relationship between DNA methylation and regulation of secondary metabolism.展开更多
[Objectives] Flow cytometry is widely used to identify plant chromosome ploidy because of its simplicity, rapidity and accuracy. Chromosome ploidy identification is an important part of sugarcane ploidy breeding and a...[Objectives] Flow cytometry is widely used to identify plant chromosome ploidy because of its simplicity, rapidity and accuracy. Chromosome ploidy identification is an important part of sugarcane ploidy breeding and application research. It is particularly important to find out the best detection part for ploidy identification in sugarcane. [Methods] The cell suspensions of sugarcane stem tips and leaves were prepared by blade chopping method. The cell suspensions were detected by flow cytometry. The best position for ploidy identification was determined by comparing the cell suspension prepared from stem tips and cell suspension prepared from leaves. [Results] The results showed that the cell suspension dissociated from stem tips was more clear than that from leaf cell suspension;the proportion of non-adherent cells in the suspension prepared from stem tips was larger than that from the leaf cell suspension;the main peak of the stem tip cell suspension was single and the number of cells was more than that of the stem tip cell suspension by flow cytometry. Using the known ploidy ‘Badilar’ as the internal reference, the ploidy of cyathomi 87-16 was detected to be 8.37. [Conclusions] Sugarcane shoot tips are an ideal material for ploidy identification. This study provides a theoretical basis for selecting the best detection site for ploidy identification of sugarcane.展开更多
文摘A microfabricated electrical impedance spectroscopy (EIS) chip with microelectrodes was developed.The substrate and the electrodes of the chip were made of glass and gold,respectively.The experimental results demonstrated that the EIS-chip could distinguish different solutions (physiological saline,culture medium,living cell suspension etc.) by scanning from 10Hz to 45kHz.A 6-element circuit model was used for fitting the real part and the imaginary part admittance curves of the living cell suspension.An actual circuit was also built and tested to verify the 6-element circuit model proposed.The micro-EIS chip has several advantages including the use of small sample volumes,high resolution and ease of operation.It shows good application prospects in the areas of cellular electrophysioiogy,drug screening and bio-sensors etc.
基金supported by NIH Core Grants P30-EY008098the Eye and Ear Foundation of Pittsburghunrestricted grants from Research to Prevent Blindness,New York,NY,USA(to KCC)。
文摘Glaucoma,characterized by a degenerative loss of retinal ganglion cells,is the second leading cause of blindness worldwide.There is currently no cure for vision loss in glaucoma because retinal ganglion cells do not regenerate and are not replaced after injury.Human stem cell-derived retinal ganglion cell transplant is a potential therapeutic strategy for retinal ganglion cell degenerative diseases.In this review,we first discuss a 2D protocol for retinal ganglion cell differentiation from human stem cell culture,including a rapid protocol that can generate retinal ganglion cells in less than two weeks and focus on their transplantation outcomes.Next,we discuss using 3D retinal organoids for retinal ganglion cell transplantation,comparing cell suspensions and clusters.This review provides insight into current knowledge on human stem cell-derived retinal ganglion cell differentiation and transplantation,with an impact on the field of regenerative medicine and especially retinal ganglion cell degenerative diseases such as glaucoma and other optic neuropathies.
文摘The effect of La, Ce was firstly tested on growth of the Taxus(T.) cuspidata cell suspensions, biosynthesis and release of taxol. The results indicate that the growth pattern of T.cuspidata cells is altered significantly by adding high concentration of rare earth in the medium. The lag and exponential phases of cell growth are shortened, the stationary phase disappears and the biomass fluctuates periodically during the decline phase. The rare earth compounds added in the exponential phase obviously increase the taxol biosyntheis and release yields of T.cuspidata cells, and the supplement of carbon source in the medium containing rare earth is also favorable to taxol biosynthesis.
基金the Tianjin Science and Technology Commission Key Project,No.07JCZDJC08000the Natural Science Foundation of China, No.30772193,30571876National High-Tech R&D Program of China (863 Program),No.2007AA04Z235
文摘Transplantation of activated transgenic Schwann cells or a fetal spinal cord cell suspension has been widely used to treat spinal cord injury. However, little is known regarding the effects of co-transplantation. In the present study, autologous Schwann cells in combination with a fetal spinal cord cell suspension were transplanted into adult Wistar rats with spinal cord injury, and newly generated axonal connections were observed ultrastructurally. Transmission electron microscopic observations showed that the neuroblast first presented cytoplasmic processes, followed by pre- and postsynaptic membranes with low electron density forming a dense projection. The number and types of synaptic vesicles were increased. Synaptic connections developed from single cell body-dendritic synapses into multiple cell body-dendritic and dendrite-dendritic synapses. In addition, the cell organs of the transplanted neuroblast, oligodendroblast and astroblast matured gradually. The blood-brain barrier appeared subsequently. Moreover, neurofilament, histamine, calcitonin-gene-related peptides, and glial fibrillary acidic protein positive fibers were observed in the transplant region. These findings demonstrate that fetal spinal cord cells in the presence of autologous activated Schwann cells can develop into mature synapses in the cavity of injured spinal cords, suggesting the possibility of information exchange through the reconstructed synapse between fetal spinal cord cells and the host.
基金This work is supported by the National Natural Science Foundation of China(to Jungui Dai,No.30100230).
文摘Deacetoxy-13-oxo sinenxan A (1) was converted to 9a-hydroxy-13-oxo-2a, 5a, 10b-triacetoxy-4(20),11-taxadiene (2) and 10b-hydroxy-13-oxo-2a,5a,9a-triacetoxy- 4(20), 11- taxadiene (3) by Ginkgo cell suspension cultures in 45% and 15% yields, respectively.
基金the National "948" Plan Project in China (Grant No.2006-4-72)
文摘Leaves of fine Populus tomentosa genotype TC152 were used as explants to establish cell suspension lines. The effects of plant growth regulators on callus induction and establishment of cell suspension lines were studied. The callus induction rate was the highest on a MS solid medium supplemented with 1.0 mg·L^-1 2,4-D. A cell suspension line could be obtained by inoculating calli which were not subcultured into a MS liquid medium supplemented with 1.5 mg·L^-1 2,4-D. The best subculture medium was MS + 0.8 mg'L-1 2,4-D + 30 g·L^-1 sucrose with a subculture cycle of seven days.
文摘A cell suspension culture of Panax ginseng which may be continuously subcultured has been established. Embryogenic callus derived from clutured young leaves was used to initiate the culture.Plant growth regulators, basal medium formula and carbohydrate levels were examined to determine their various effects on suspension culture cell growth and development. The best selection of plant growth regulator, hasal medium and carbohydrate level is 2 mg / L 2,4-D: 0.5 mg / L KT,MS and 3% sucrose respectively.
文摘The standard detection hallmarks of apoptosis of Taxus cuspidata cells in suspension culture with Ce 4+ were studied. The condensation and margination of chromatin were observed under the electron microscopy. DNA fragmentation ranged 'DNA ladder' on agarose gel electrophoresis. TdT mediated dUTP nick end labeling (TUNEL) analysis of the cells reveals that the nuclear DNA strand breaks can be identified by labeling free 3′ OH termini. These results suggest that Ce 4+ can induce apoptosis of Taxus cuspidata cells and also indicate that there is a certain relationship between apoptosis and secondary metabolite product Taxol.
文摘Suspension cultures initiated from callus derived from petiole explants of aspen hybrid (Populus tremuloides×P. tremula) produced somatic embryos. Callus was induced on a MS medium supplemented with 5mg·L^-1 2,4-D and 0.05mg·L^-1 zeatin under light conditions. Embryogenic calli were obtained when a subsequent subculture of calli was suspended in the same basal medium with 10mg·L^-1 2,4-D. The highest number of globular embryos were induced from embryogenic calli by cell suspension culture in a MS liquid medium supplemented with 10mg·L^-1 2,4-D. Genotype and 2,4-D concentration were vital to the induction of embryogenic calli producing competent cells. Embryogenic calli for each genotype were heterogeneous. Green calli with gel-like consistency could yield more competent cells than light yellow embryogenic calli. However, some globular embryos broke into slices and some developed abnormally after one month of culture under the same or other hormonal conditions.
基金supported by the National Key Research and Development Program of China(2016YFD0600106)the National Natural Science Foundation of China(31670675)the Fundamental Research Funds for the Central Universities(2572018CL01 and 2572018CL02).
文摘Populus spp.have long been used as model woody plant species for molecular biology research.However,tissues of poplar are often recalcitrant to experimental procedures for molecular studies.We generated a hormone autotrophic poplar suspension cell line from a hybrid of Populus alba×P.berolinensis‘Yinzhong’,named Qu-2.Qu-2 cells are suitable as a model biological system for studying woody plants.Qu-2 cells have many advantages over suspension cell lines derived so far from any other woody plants.Qu-2 cells are very easy to cultivate and can grow on several common plant culture media without the addition of any plant hormone.They show exceptionally high growth rates,reaching an approximately 150-fold increase in biomass after one week of culturing.Another important unique characteristic of Qu-2 cells is that they can be cryopreserved and readily reactivated.Qu-2 cells are suitable for molecular manipulations such as protoplast production,transient transformation,and RNA-seq analysis.Therefore,Qu-2 cells have the great potential to be an excellent model cell line in tree molecular biological research,ranging from physiology to gene function.The Qu-2 cells will be made available to the plant community for research.
文摘Friable callus was induced when immature seeds of floribunda rose were inoculated on MS medium supplemented with 2,4-D 3.0 mg.L^-1. When transfered onto subculture media, friable callus developed into embryogenic callus, which was used to establish cell suspension lines. Cell suspensions had to be subcultured at a interval of 4-5 days at the first several culture cycles. The best subculturing cycle for the stable cell suspensions was 8-10 days. The best inoculum quantity was 1 mL PCV(Packed Cell Volume) per 40 mL culture fluid.
文摘The activity of alcohol dehydrogenase (ADH) in cultured cells of various tobacco was determined. It was found that significant differences existed in cells of different varieties cultured under normal conditions and as well after treated with exogenous ethanol. The ADH activity had positive relation with the ability of the cells to catabolize exogenous ethanol, indicating that the main function of the ADH in tobacco cells was in the direction of converting ethanol to acetaldehyde.
文摘In this work, we present an ultrastructural and physiological description of a novel chlorophyll-deficient, yellow cell line of the grass Bouteloua gracilis that develops etioplast-like plastids in presence of light (YELP). These mutant cells were compared to the parental, wild-type, highly chlorophyllous cells from which they were isolated. Growth analysis, based on fresh and dry weights, indicated that YELP accumulates biomass at a slower rate than the parental, green cells. Besides, YELP developed very low levels of photosynthetic pigments, reaching only 9.3% and 38.4% of chlorophyll a and chlorophyll b, respectively, developed by the wild-type cells. Likewise, the accessory pigments, carotenes and xanthophylls, were only synthesized at 8.0% and 5.4%, respectively, of the levels reached by the green cells. Electron microscopy revealed remarkable differences in plastid ultrastructure between the wild-type and mutant cells. Plastids of YELP were heterogeneous and smaller than those found in wild-type cells. YELP plastids were abnormal with poorly developed membrane systems that prevented the accumulation of chlorophyll and accessory pigments in the mutant cell line. We expect this novel, mutant cell line will provide new tools for studying plastid development and differentiation.
文摘We used the leaf blade of rice (cultivars were Nonghu 6, Sugeng 2, Huyou 2 and Hanfeng) as initial material for protoplast culture, and a great number of regenerated plants were obtained. Rice seeds were sterilized and germinated. The immature leaves were cut into 3-5 mm pieces when the third or forth leaf appeared. Leaf pieces were inoculated on MS medium with 2,4-D 4 mg/1, NAA 2mg/1 and IAA Img/1. After 2 wk culture, calli were induced and subcultured once or twice for multiplication. 3-5 g calli were transferred to the modified MS liquid medium with 2,4-D 2 mg/1 and KT 0.5mg/1 for suspension culture. Embryogenic cell suspension was established after 2 mo culture. The effect of the growth period of suspension cells on the
文摘In vitro tissue culture of hard woody, endangered, medicinal plant Coscinium fenestratum is most challenging to plant tissue culturists. In the present study, petiole and leaf explants of Coscinium fenestratum were induced to form callus when cultured on vermicompost extract media along with coelomic fluid. Suspension medium was developed using vermicompost extract and coelomic fluid in 3:1 ratio. Phytochemical analysis of the alkaloid berberine was confirmed from callus, suspension cell culture and suspension medium by Thin Layer Chromatography and High Performance Liquid Chromatography. Vermicompost and its extracts with coelomic fluid have shown maximum (100 per cent) response of callus induction. Callus mass enlarged with increasing concentration of coelomic fluid and callus growth was assessed from the biomass. Incubation of culture tubes in dark supported callus development significantly. The Rf value of 0.36 confirmed the presence of berberine by Thin Layer Chromatography. Qualitative analysis confirmed the presence of alkaloid berberine with the retention time of 2.8 minutes similar to that of standard reference sample from Sigma chemicals, USA. The suspension medium turned deep yellow because of the release of the alkaloid. Vermicompost and its extracts along with coelomic fluid have shown the economical approach for micropropagation of economically and medicinally important plants.
基金Supported by General Project of National Natural Science Foundation of China(41371470)Key Program of Natural Science Foundation of Hubei Province(2011CDA037)Science and Technology Planning Project of Huangshi City(2010A1019-2)
文摘Ricinus communis L. is a new copper hyperaccumulator growing on Tonglushan copper mine in Hubei Province, China. This study aimed to establish a suspension cell line of R. communis L. with stable and rapid growth for further screening of heavy metal-resistant R. communis L. cells and breeding of hyperaccu- mulators. In this study, cell suspension culture conditions were optimized by using orthogonal experimental design with previously induced R. communis L. embyre- genic calluses as experimental materials, to establish the suspension cell line of R. communis L. Under the optimal conditions, growth curves of suspension cells and changes in pH of culture liquid were determined. The results showed that the optimal culture conditions for R. conmmnis L. suspension cell line were : MS + O. 5 rag/I, 6-BA + O. 2 mg/L NAA + 50 mg/L sucrose + 350 mg/L casein hydrolysate as basic medium, with callus inoculation amount of 2.5 g/50 ml, dark culture at (26 ±2) ℃ with shaking at 110 r/min. Under these conditions, increment of fresh weight and dry weight ofR. commun/s L. suspension cells reached the maximum of 4.56 g/(50 ml 14 d) and 0.49 g/(50 ml 16 d), respectively. Growth curves of R. communis L. suspension cells were basically in "S" shape. Each culture cycle lasted 16 d, and the rapid growth stage was from the 6th d to the 14th d. In a culture cycle, pH of the culture liquid declined first and then increased to the maximum and stabilized gradually.
基金This research was funded by the Provincial Supporting Program of Hebei for the Returned Oversea Scholars,China(C20210114)the Science and Technology Project of Hebei Education Department(QN2022017)+5 种基金the Fundamental Scientific Research Fund of Universities in Hebei Province(KY2021059)the China Agriculture Research System(CARS-30-2-07)the National Key Research and Development Project of China(2019YFD1001605)the Natural Science Foundation of Hebei Province(C2020204082)the Funds for Hebei Jujube Industry Technology Research Institute after Operation Performance(205676155H)the Young Talent Project of Hebei Agricultural University Foundation(YJ201853).
文摘3´,5´-Cyclic adenosine monophosphate(cAMP)is an important metabolite that is specifically enriched in jujube.However,the effect of cAMP on jujube cellular responses has not been comprehensively studied.Here,we established jujube cell suspension cultures and investigated the calcium influx in response to cAMP treatment through protoplast isolation and fluorescence intensity.Firstly,cAMP treatment could promote jujube growth and increase the content of endogenous cAMP.Using transcriptome analysis with transgenic Arabidopsis plants overexpressing adenylate cyclase(ZjAC)as a positive control,we identified 60 calcium-related differential expressed genes(DEGs)that contributed to the calcium signaling and inter-or intra-cellular responses.Pharmacological treatments such as cAMP and the calcium ionophore A23187 could induce ZjAC expression,the accumulation of cAMP and calcium influx in jujube cells,while ethylene glycol tetraacetic acid(EGTA)or bithionol treatment inhibited these changes.Moreover,the calcium channels and transporters in calcium influx,such as the ZjCNGC2 channel and the mitogen activated protein(MAP)kinase pathway,could be activated by cAMP treatment.In summary,our findings demonstrated that cAMP biosynthesis is dependent on calcium influx and the amplifying effect between calcium and cAMP may be involved in intracellular signal induction,which might contribute to the growth and development of jujube.
文摘The authors used suspension cells of Populus euphratica to isolate protoplast in the present study. Protoplasts were successfully obtained after 4 hours incubation in enzyme solution containing 1 0% cellulase “onozuka” R\|10, 0\^01% pectolyase Y\|23,0\^15% macerozyme R\|10 and 0\^1% hemicellulase at 25℃. Outward and inward single channels in plasma membrane were observed using cell\|attached recording of patch\|clamp technique. In this study, single channel records showed that more than one species of channel were obtained. These attempts in protoplast isolation and ion channel recording offers the opportunity to characterize cellular mechanisms of salt tolerance in tree species.
基金The work was supported by The Fundamental Research Funds for the Central Universities(2572017EA05)the National Natural Science Foundation of China(31870588).
文摘DNA methylation is widespread in plants and associated with plant development and defense mechanisms.However,the relationship between DNA methylation and plant secondary metabolism has rarely been reported.Here,when birch suspension cells were treated with 5-azacytidine(5-azaC),which blocks DNA methylation,triterpenoid accumulation was significantly promoted and antioxidant and defense enzymatic activity changed.For studying triterpenoid accumulation,0.1 mM azaC was optimal.A qRT-PCR assay revealed increased expression of genes encoding key triterpenoid biosynthetic enzymes.Evaluation of methylation polymorphisms at CCGG sites showed that the methylation level was lower in cells treated with 5-azaC.These results demonstrated that 5-azaC treatment led to an increase in the production of triterpenoids in cell cultures through a mechanism that involved in DNA methylation,which resulted in the induction of genes encoding the key enzymes.The study provides evidence of a relationship between DNA methylation and regulation of secondary metabolism.
基金Supported by National Natural Science Foundation of China(31860350)Special Fund for Basal Scientific Research of Guangxi Academy of Agricultural Sciences(GNK 2020YM24)Science and Technology Development Fund of Guangxi Academy of Agricultural Sciences(2021YT006)。
文摘[Objectives] Flow cytometry is widely used to identify plant chromosome ploidy because of its simplicity, rapidity and accuracy. Chromosome ploidy identification is an important part of sugarcane ploidy breeding and application research. It is particularly important to find out the best detection part for ploidy identification in sugarcane. [Methods] The cell suspensions of sugarcane stem tips and leaves were prepared by blade chopping method. The cell suspensions were detected by flow cytometry. The best position for ploidy identification was determined by comparing the cell suspension prepared from stem tips and cell suspension prepared from leaves. [Results] The results showed that the cell suspension dissociated from stem tips was more clear than that from leaf cell suspension;the proportion of non-adherent cells in the suspension prepared from stem tips was larger than that from the leaf cell suspension;the main peak of the stem tip cell suspension was single and the number of cells was more than that of the stem tip cell suspension by flow cytometry. Using the known ploidy ‘Badilar’ as the internal reference, the ploidy of cyathomi 87-16 was detected to be 8.37. [Conclusions] Sugarcane shoot tips are an ideal material for ploidy identification. This study provides a theoretical basis for selecting the best detection site for ploidy identification of sugarcane.