Agrobacterium tumefaciens-mediated transformation has been widely adopted for plant genetic engineering and the study of gene function(Krenek et al.,2015).This method is prevalent in the genetic transformation of herb...Agrobacterium tumefaciens-mediated transformation has been widely adopted for plant genetic engineering and the study of gene function(Krenek et al.,2015).This method is prevalent in the genetic transformation of herbaceous plants,with notable applications in species such as Arabidopsis(Yin et al.,2024),soybean(Zhang et al.,2024),rice(Zhang et al.,2020),and Chinese cabbage(Li et al.,2021).However,its application in fruit trees is limited.This is primarily due to their long growth cycles and lack of rapid,efficient,and stable transgenic systems,which severely hinders foundational research involving plant genetic transformation(Mei et al.,2024).Furthermore,for subtropical fruit trees,the presence of recalcitrant seeds adds an extra layer of difficulty to genetic transformation(Umarani et al.,2015),as most methods rely on seed germination as a basis for transformation.展开更多
Cannabis sativa is highly valued for its use in fiber production,medicine,and recreational products.Its secondary metabolites(SM)are renowned for their wide range of health benefits and psychoactive properties.While m...Cannabis sativa is highly valued for its use in fiber production,medicine,and recreational products.Its secondary metabolites(SM)are renowned for their wide range of health benefits and psychoactive properties.While much of the existing research has focused on cannabinoid production in the plant’s aerial parts,particularly the leaves and flowers,the root system remains understudied in terms of its SM profile.One promising in vitro approach for metabolite production involves the use of‘hairy roots(HRs)’.These roots mimic the phytochemical profile of native roots but grow more efficiently and yield higher quantities of metabolites.HRs are genetically altered root tissues that develop at the site of infection when Agrobacterium rhizogenes is introduced into wounded plant tissues.HRs cultures in Cannabis represent a breakthrough in plant metabolic engineering,offering potential for the controlled biosynthesis of cannabinoids and terpenoids.By utilising genome editing(GE)tools such as CRISPR-based tools,these cultures can produce novel bioactive compounds at an industrial scale.The use of elicitors enhances the production of SM by activating their biosynthetic pathways,further boosting yields.This system provides a sustainable alternative to conventional farming and chemical synthesis,addressing challenges such as pharmaceutical shortages,enhancing climate resilience,and promoting more resource-efficient biomanufacturing.Few studies have explored elicitor-induced HR cultures in Cannabis to enhance terpenoid production.This review highlights research on HRs for SM synthesis and introduces a platform that positions Cannabis as a leader in biomanufacturing and sustainable biotechnology,promoting advancements across the agricultural and pharmaceutical industries globally.展开更多
In this work,we apply tunneling formalism to analyze charged particles tunneling across a hairy black hole horizon.Such black hole solutions are essential for frameworks based on Horndeski's gravity theory.Applyin...In this work,we apply tunneling formalism to analyze charged particles tunneling across a hairy black hole horizon.Such black hole solutions are essential for frameworks based on Horndeski's gravity theory.Applying a semi-classical technique,we examine the tunneling of charged particles from a hairy black hole and derive the generic tunneling spectrum of released particles,ignoring self-gravitational and interaction.It is studied to ignore the back-reaction impact of the radiated particle on the hairy black hole.We analyze the properties of the black hole,such as temperature and entropy,under the influence of quantum gravity and also observe that the firstorder correction is present.We study tunneling radiation produced by a charged field equation in the presence of a generalized uncertainty effect.We modify the semi-classical technique by using the generalized uncertainty principle,the WKB approximation,and surface gravity.展开更多
The CRISPR/Cas9 genome-editing system serves as a pivotal tool for enhancing crop genetics.Within this system,single guide RNAs(sgRNAs)are instrumental in the precise cleavage of DNA double strands.However,the efficie...The CRISPR/Cas9 genome-editing system serves as a pivotal tool for enhancing crop genetics.Within this system,single guide RNAs(sgRNAs)are instrumental in the precise cleavage of DNA double strands.However,the efficiency of gene editing varies among sgRNAs,emphasizing the need to meticulously select target sites,especially in the context of Citrullus lanatus,a species notorious for its challenging and inefficient generation of transgenic plants through stable transformation.This study employed an Agrobacterium rhizogenes-mediated hairy root method to assess effective target sites for gene editing of ClCIPK17 across various Citrullus species.Hairy roots were successfully induced in different plant tissues at diverse growth stages of Citrullus lanatus,Citrullus mucosospermus,and Citrullus amarus.Employing a vector with two sgRNAs(sgRNA1 and sgRNA5)positioned within conserved regions of exon 1 and exon 5 of ClCIPK17 in the CRISPR/Cas9 system,targeted mutations were detected in 90.9%of accessions across the four Citrullus species.Notably,73.94%of all examined hairy roots exhibited mutations at the sgRNA1 site,while the sgRNA5 site showed no mutations.Among the 31 different mutation types identified at the sgRNA1 site,base deletion was the most prevalent.Using the sgRNA1 site of ClCIPK17,stable transgenic watermelon buds were obtained from explants,and the targeted mutations of the sgRNA1 site were confirmed.These findings underscore the viability of the hairy root transformation system in assessing the editing efficiency of sgRNA targets in diverse Citrullus species.展开更多
The artemisinin accumulation in the hairy root cultures of Artemisia annua L. was enhanced via a treatment of three fungal elicitors separately ( Verticillium dahliae Kleb., Rhizopus stolonifer (Ehrenb. ex ...The artemisinin accumulation in the hairy root cultures of Artemisia annua L. was enhanced via a treatment of three fungal elicitors separately ( Verticillium dahliae Kleb., Rhizopus stolonifer (Ehrenb. ex Fr.) Vuill and Colletotrichum dematium (Pers.) Grove). Among these three elicitors, V. dahliae had the highest inducing efficiency, but none of them manifests any noticeable effects on the cell growth of the hairy root cultures. The artemisinin content of the hairy root cultures treated with V. dahliae elicitor was 1.12 mg/g DW, which was 45% higher than the control (0.77 mg/g DW). The results showed that elicitation was dependent on the elicitor concentration, the incubation period and the physiological stage at which the hairy root cultures were treated. In addition, the authors found that for V. dahliae , the optimum concentration was 0.4 mg carbohydrate per millilitre medium, the strongest response of A. annua hairy root cultures to the elicitation was at the late exponential growth stage, and the highest artemisinin content of the hairy root cultures was on the 4th day post treatment.展开更多
The biotransformation of artemisinin by hairy root cultures ofRheum palmatum L. was investigated for the first time. The main product, deoxyartemisinin, was isolated and characterized on the basis of its spectral data.
The Hairy Ape is a representative play of Eugene O'Neill that distinguishes collective trauma of mechanization and materialism in the 20 th century of America. Till now, there have not been any thesis discussing T...The Hairy Ape is a representative play of Eugene O'Neill that distinguishes collective trauma of mechanization and materialism in the 20 th century of America. Till now, there have not been any thesis discussing The Hairy Ape from the perspective of trauma theory. And the studies of other works on trauma theory is also not many. Based on trauma theory, this thesis makes a decoding in The Hairy Ape.展开更多
The NAC （NAM, ATAF1/2 and CUC2） transcription factor family plays a key role in plant development and responses to abiotic stress. GmNAC15 （Glyma 15g40510.1）, a member of the NAC transcription factor family in soy...The NAC （NAM, ATAF1/2 and CUC2） transcription factor family plays a key role in plant development and responses to abiotic stress. GmNAC15 （Glyma 15g40510.1）, a member of the NAC transcription factor family in soybean, was functionally characterized, especially with regard to its role in salt tolerance. In the present study, qRT-PCR （quantitative reverse transcription PCR） analysis indicated that GmNAC15 was induced by salt, drought, low temperature stress, and ABA treatment in roots and leaves. GmNAC15 overexpression in soybean （Glycine max） hairy roots enhanced salt tolerance. Transgenic hairy roots improved the survival of wild leaves; however, overexpression of GmNAC15 in hairy root couldn＇t influnce the expression level of GmNAC15 in leaf. GmNAC15 regulates the expression levels of genes responsive to salt stress. Altogether, these results provide experimental evidence of the positive effect of GmNAC15 on salt tolerance in soybean and the potential application of genetic manipulation to enhance the salt tolerance of important crops.展开更多
Black hairy tongue (BHT) is a benign medical condition characterized by elongated filiform lingual papillae with typical carpet-like appearance of the dorsum of the tongue. Its prevalence varies geographically, typica...Black hairy tongue (BHT) is a benign medical condition characterized by elongated filiform lingual papillae with typical carpet-like appearance of the dorsum of the tongue. Its prevalence varies geographically, typically ranging from 0.6% to 11.3%. Known predisposing factors include smoking, excessive coffee/black tea consumption, poor oral hygiene, trigeminal neuralgia, general debilitation, xerostomia, and medication use. Clinical presentation varies but is typically asymptomatic, although aesthetic concerns are common. Differential diagnosis includes pseudo-BHT, acanthosis nigricans, oral hairy leukoplakia, pigmented fungiform papillae of the tongue, and congenital melanocytic/melanotic nevi/macules. Clinical diagnosis relies on visual observation, detailed history taking, and occasionally microscopic evaluation. Treatment involves identification and discontinuation of the offending agent, modifications of chronic predisposing factors, patient’s re-assurance to the benign nature of the condition, and maintenance of adequate oral hygiene with gentle debridement to promote desquamation. Complications of BHT (burning mouth syndrome, halitosis, nausea, gagging, dysgeusia) typically respond to therapy. Prognosis is excellent with treatment of underlying medical conditions. BHT remains an important medical condition which may result in additional burden on the patient and health care system and requires appropriate prevention, recognition and treatment.展开更多
Catharanthus roseus contains important anti-tumor terpenoid indole alkaloids (TIAs) such as vinblastine and vincristine. Cytochrome P450 enzyme geraniol 10-hydroxylase (G10H) is a putative rate-limiting enzyme involve...Catharanthus roseus contains important anti-tumor terpenoid indole alkaloids (TIAs) such as vinblastine and vincristine. Cytochrome P450 enzyme geraniol 10-hydroxylase (G10H) is a putative rate-limiting enzyme involved in the TIAs biosynthetic pathway in C. roseus. In this study the g10h gene driven by the cauliflower mosaic virus 35S (CaMV 35S) promoter was introduced into C. roseus through Agrobacterium-mediated transformation. The integration and overexpression of the target gene (g10h) in hairy root lines were confirmed by polymerase chain reaction and RT-QPCR analysis respectively. Overexpression of g10h in transgenic hairy root lines significantly enhanced the accumulations of monomeric alkaloid ajmalicine and dimeric alkaloids, vincristine and vinblastine. Total TIAs production in hairy roots reached (9.51) mg/g DW, over 3-fold higher than that in the untransformed root lines. This is the first report that engineering of g10h into TIAs-producing plant species results in significant enhancement of TIAs accumulation in cultured hairy roots. This study demonstrates that the putative rate-limiting step catalyzed by G10H is indeed the real rate-limiting step involved in the TIAs biosynthetic pathway in C. roseus, which is one of the key targets for promoting TIAs production by genetic engineering.展开更多
Objective:To increase biomass and saponin production in hairy root culture of Talinum paniculatum Gaertn.(T.paniculatum)in balloon-type bubble bioreactor(BTBB).Methods:Hairy roots which were collected from leaf explan...Objective:To increase biomass and saponin production in hairy root culture of Talinum paniculatum Gaertn.(T.paniculatum)in balloon-type bubble bioreactor(BTBB).Methods:Hairy roots which were collected from leaf explants of T.paniculatum were infected by Agrobacterium rhizogenes strain LB510.The hairy roots were cultivated at400 m L Murashige and Skoog liquid medium without growth regulator(MS0)in1 000 m L BTBB.Each BTBB had 2 g hairy roots as initial inoculum and these cultures were treated with various concentrations of sucrose(3%,4%,5%,6%w/v)and potassium nitrate(0.5,1.0,1.5 and 2.0 strength of MS medium).Cultures were maintained for 14days.Fresh and dry weights of hairy roots at the end of culture were investigated.Results:Various concentrations of sucrose influenced the biomass accumulation of hairy roots.Maximum biomass was reached by MS medium supplemented with 6%sucrose and it was approximately threefold higher than control.Culture supplemented with potassium nitrate at 2.0 strength of MS0 could increase biomass accumulation of hairy roots until 0.14 g dry weight and it was almost threefold higher than control.However,the maximum saponin content was obtained by MS medium supplemented with 5%sucrose and 2.0 strength potassium nitrate of MS.Conclusions:Based on this research,those conditions can be used to produce biomass and saponin of hairy root of T.paniculatum in the large scale.展开更多
Glycogen synthase kinase 3(GSK3)is a kind of sedne/threonine kinase widely found in eukaryotes.Many plant GSK3 kinases play important roles in regulating stress responses.This study investigated BRASSINOSTEROID-INSENS...Glycogen synthase kinase 3(GSK3)is a kind of sedne/threonine kinase widely found in eukaryotes.Many plant GSK3 kinases play important roles in regulating stress responses.This study investigated BRASSINOSTEROID-INSENSITIVE 2(GmBIN2)gene,a member of the GSK3 protein kinase family in soybean and an orthologue of Arabidopsis BIN2/AtSK21.GmBIN2 expression was increased by salt and drought stresses,but was not significantly affected by the ABA treatment.To examine the function of GrnBIN2,transgenic Arabidopsis and transgenic soybean hairy roots were generated.Overexpression of GmBIN2 in Arabidopsis resulted in increased germination rate and root length compared with wild-type plants under salt and mannitol treatments.Overexpression of GmBIN2 increased cellular Ca2~content and reduced Na~content,enhancing salt tolerance in transgenic Arabidopsis plants.In the soybean hairy root assay,overexpression of GmBIN2 in transgenic roots also showed significantly higher relative root growth rate than the control when subjected to salt and mannitol treatments.Measurement of physiological indicators,including proline content,superoxide dismutase(SOD)activity,and relative electrical conductivity,supported this conclusion.Furthermore,we also found that GmBIN2 could up-regulate the expression of some stress-related genes in transgenic Arabidopsis and soybean hairy roots.Overall,these results indicated that GmBIN2 improved tolerance to salt and drought in transgenic Arabidopsis and soybean hairy roots.展开更多
Soybean is one of the world's most important oil and protein crops. Efficient transformation is a key factor for the improvement of soybean by genetic modification. We describe an optimized protocol for the Agroba...Soybean is one of the world's most important oil and protein crops. Efficient transformation is a key factor for the improvement of soybean by genetic modification. We describe an optimized protocol for the Agrobacterium rhizogenes-mediated transformation of soybean and the induction of hairy root development in vitro. Cotyledons with 0.5-cm hypocotyls were cut from 5-day-old seedlings and used as explants. After infection and co-cultivation,hairy roots were produced in induction culture medium after 10–12 days. Using this method, 90%–99% of the infected explants of five different cultivars produced hairy roots within one month. Observations using reporter constructs showed that 30%–60% of the hairy roots induced were transformed. Based on high transformation efficiency and short transformation period, this method represents an efficient and rapid platform for study of soybean gene function.展开更多
The glycosylation of hydroxylcoumarin was investigated by using suspension cultures of hairy roots of Polygonum multiflorum. Two new coumarin glucosides （3 and 4） were biosysthesized by regioselectively glycosylatio...The glycosylation of hydroxylcoumarin was investigated by using suspension cultures of hairy roots of Polygonum multiflorum. Two new coumarin glucosides （3 and 4） were biosysthesized by regioselectively glycosylation of corresponding substrates （1 and 2） in the system. The structures of two products were identified as 7-hydroxy-4-methylcoumarin 5-O-β-D-glucopyranoside and 7- hydroxy-3,4-dimethylcoumarin 5-O-β-D-glucopyranoside on the ground of chemical and spectroscopic methods, respectively. 2007 Rong Min Yu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.展开更多
The legume forage Alhagi pseudoalhagi was transformed by the Agrobacterium rhizogenes strain A4 using cotyledon and hypocotyl segments as infection materials. Regenerated plants were achieved from sterile calli derive...The legume forage Alhagi pseudoalhagi was transformed by the Agrobacterium rhizogenes strain A4 using cotyledon and hypocotyl segments as infection materials. Regenerated plants were achieved from sterile calli derived from hairy roots, which occurred at or near the infection sites. The regenerated plants from hairy root were characterized by normal leaf morphology and stem growth but a shallow and more extensive root system than normal plants. Opine synthesis, PCR and Southern blot confirmed that T- DNA had been integrated into the A. pseudoalhagi genome. Acetosyringone (AS) was found to be vital for successful transformation of A. pseudoalhagi.展开更多
The effects of various rare earth elements on growth and isoflavonoid production in hairy root cultures of Pueraria lobata (Willd.) Ohwi (strain TR2) cultured in 500 ml flasks were studied. After 32 days of culture, t...The effects of various rare earth elements on growth and isoflavonoid production in hairy root cultures of Pueraria lobata (Willd.) Ohwi (strain TR2) cultured in 500 ml flasks were studied. After 32 days of culture, the biomass of hairy roots increase 15 times and reach 3.2 g dry weight. Hairy root growth was inhibited by Y_2O_3, NaSeO_3 and Sm^(3+) because of brown formation. But significant promoting effect on root growth due to callus formation was observed in La^(3+) treatments. It is the most noteworthy that the production of total isoflavonoids and puerarin was enhanced greatly by La^(3+) treatment. A major portion of increased total isoflavonoids and puerarin was released into medium in La^(3+) treatment while the hairy root viabilities were preserved. Some specific secondary metabolite release processes could be induced by La^(3+) and their possible mechanism is discussed.展开更多
Muscle-derived stem cells were isolated from the skeletal muscle of Sprague-Dawley neonatal rats aged 3 days old. Cells at passage 5 were incubated in Dulbecco＇s modified Eagle＇s medium supplemented with 10% （v/v）...Muscle-derived stem cells were isolated from the skeletal muscle of Sprague-Dawley neonatal rats aged 3 days old. Cells at passage 5 were incubated in Dulbecco＇s modified Eagle＇s medium supplemented with 10% （v/v） fetal bovine serum, 20 IJg/L nerve growth factor, 20 pg/L basic fibroblast growth factor and 1% （v/v） penicillin for 6 days. Cells presented with long processes, similar to nerve cells. Connections were formed between cell processes. Immunocytochemical staining with neuron specific enolase verified that cells differentiated into neuron-like cells. Immunofluorescence cytochemistry and western blot results revealed that the expression of protein hairy enhancer of split-1 was significantly reduced. These results indicate that low expression of protein hairy enhancer of split-1 participates in the differentiation of muscle-derived stem cells into neuron-like cells.展开更多
We present some novel rotating hairy black hole metric in （2 ＋ 1） dimensions, which is an exact solution to the field equations of the Einstein-scalar-AdS theory with a non-minimal coupling. The scalar potential is...We present some novel rotating hairy black hole metric in （2 ＋ 1） dimensions, which is an exact solution to the field equations of the Einstein-scalar-AdS theory with a non-minimal coupling. The scalar potential is determined by the metric ansatz and consistency of the field equations and cannot be prescribed arbitrarily. In the simplified, critical ease, the scalar potential contains two independent constant parameters, which are respectively related to the mass and angular momentum of the black hole in a particular way. As long as the angular momentum does not vanish, the metric can have zero, one or two horizons. The case with no horizon is physically uninteresting because of the curvature singularity lying at the origin. We identify the necessary conditions for at least one horizon to be present in the solution, which imposes some bound on the mass-angular momentum ratio. For some particular choice of pararneters our solution degenerates into some previously known black hole solutions.展开更多
Tetrahydrocannabinol (THC) derivatives are used clinically as analgesic, anti-inflammatory, appetite stimulant, anti-emetic and anti-tumor cannabinoids. THC and its related compounds are at present obtained by extract...Tetrahydrocannabinol (THC) derivatives are used clinically as analgesic, anti-inflammatory, appetite stimulant, anti-emetic and anti-tumor cannabinoids. THC and its related compounds are at present obtained by extraction from intact Cannabis plants or chemical synthesis, but plant cell cultures may be an alternative source of production. In the present study, hairy root cultures of C. sativa (Cannabaceae) were induced by incubation of aseptically grown callus culture with solid B5 medium supplemented with 4 mg/l naphthaleneacetic acid in darkness at 25°C. Hairy root growth profiles in shake flask, increased periodically during 35 days of growth cycle. The cannabinoid contents produced in minor levels and remained below 2.0 μg/g dry weight. The contents of can-nabinoid were analyzed by liquid chromatography and confirmed by mass spectrometry.展开更多
基金funded by the Key-Area Research and Development Program of Guangdong Province(Grant No.2022B0202070002)the Guangxi Science and Technology Major Program(Grant No.GuikeAA23023007-2)+1 种基金the Guangdong Province Modern Agricultural Industry Technology System Innovation Team Construction Project(2024CXTD19)Guangdong Basic and Applied Basic Research Foundation(Grant No.2023A1515010303)。
文摘Agrobacterium tumefaciens-mediated transformation has been widely adopted for plant genetic engineering and the study of gene function(Krenek et al.,2015).This method is prevalent in the genetic transformation of herbaceous plants,with notable applications in species such as Arabidopsis(Yin et al.,2024),soybean(Zhang et al.,2024),rice(Zhang et al.,2020),and Chinese cabbage(Li et al.,2021).However,its application in fruit trees is limited.This is primarily due to their long growth cycles and lack of rapid,efficient,and stable transgenic systems,which severely hinders foundational research involving plant genetic transformation(Mei et al.,2024).Furthermore,for subtropical fruit trees,the presence of recalcitrant seeds adds an extra layer of difficulty to genetic transformation(Umarani et al.,2015),as most methods rely on seed germination as a basis for transformation.
基金supported by a research grant of Gyeongsangbuk-do(No.GBHEMP202504),Republic of Korea.
文摘Cannabis sativa is highly valued for its use in fiber production,medicine,and recreational products.Its secondary metabolites(SM)are renowned for their wide range of health benefits and psychoactive properties.While much of the existing research has focused on cannabinoid production in the plant’s aerial parts,particularly the leaves and flowers,the root system remains understudied in terms of its SM profile.One promising in vitro approach for metabolite production involves the use of‘hairy roots(HRs)’.These roots mimic the phytochemical profile of native roots but grow more efficiently and yield higher quantities of metabolites.HRs are genetically altered root tissues that develop at the site of infection when Agrobacterium rhizogenes is introduced into wounded plant tissues.HRs cultures in Cannabis represent a breakthrough in plant metabolic engineering,offering potential for the controlled biosynthesis of cannabinoids and terpenoids.By utilising genome editing(GE)tools such as CRISPR-based tools,these cultures can produce novel bioactive compounds at an industrial scale.The use of elicitors enhances the production of SM by activating their biosynthetic pathways,further boosting yields.This system provides a sustainable alternative to conventional farming and chemical synthesis,addressing challenges such as pharmaceutical shortages,enhancing climate resilience,and promoting more resource-efficient biomanufacturing.Few studies have explored elicitor-induced HR cultures in Cannabis to enhance terpenoid production.This review highlights research on HRs for SM synthesis and introduces a platform that positions Cannabis as a leader in biomanufacturing and sustainable biotechnology,promoting advancements across the agricultural and pharmaceutical industries globally.
基金funded by the National Natural Science Foundation of China under Grant No.11975145。
文摘In this work,we apply tunneling formalism to analyze charged particles tunneling across a hairy black hole horizon.Such black hole solutions are essential for frameworks based on Horndeski's gravity theory.Applying a semi-classical technique,we examine the tunneling of charged particles from a hairy black hole and derive the generic tunneling spectrum of released particles,ignoring self-gravitational and interaction.It is studied to ignore the back-reaction impact of the radiated particle on the hairy black hole.We analyze the properties of the black hole,such as temperature and entropy,under the influence of quantum gravity and also observe that the firstorder correction is present.We study tunneling radiation produced by a charged field equation in the presence of a generalized uncertainty effect.We modify the semi-classical technique by using the generalized uncertainty principle,the WKB approximation,and surface gravity.
基金supported by the Basic Research Funds of Chinese Academy of Agricultural Sciences(Grant No.1610192023201)the National Key Research and Development Program of China(Grant No.SQ2023YFE0201239)+3 种基金China Agriculture Research System of MOF and MARA(Grant No.CARS-25-03)the Major Science and Technology Projects of Henan Province(Grant No.221100110400)the Funding of Joint Research on Agricultural Variety Improvement of Henan Province(Grant No.2022010503)the Agricultural Science and Technology Innovation Program(Grant No.CAAS-ASTIP-2021-ZFRI)。
文摘The CRISPR/Cas9 genome-editing system serves as a pivotal tool for enhancing crop genetics.Within this system,single guide RNAs(sgRNAs)are instrumental in the precise cleavage of DNA double strands.However,the efficiency of gene editing varies among sgRNAs,emphasizing the need to meticulously select target sites,especially in the context of Citrullus lanatus,a species notorious for its challenging and inefficient generation of transgenic plants through stable transformation.This study employed an Agrobacterium rhizogenes-mediated hairy root method to assess effective target sites for gene editing of ClCIPK17 across various Citrullus species.Hairy roots were successfully induced in different plant tissues at diverse growth stages of Citrullus lanatus,Citrullus mucosospermus,and Citrullus amarus.Employing a vector with two sgRNAs(sgRNA1 and sgRNA5)positioned within conserved regions of exon 1 and exon 5 of ClCIPK17 in the CRISPR/Cas9 system,targeted mutations were detected in 90.9%of accessions across the four Citrullus species.Notably,73.94%of all examined hairy roots exhibited mutations at the sgRNA1 site,while the sgRNA5 site showed no mutations.Among the 31 different mutation types identified at the sgRNA1 site,base deletion was the most prevalent.Using the sgRNA1 site of ClCIPK17,stable transgenic watermelon buds were obtained from explants,and the targeted mutations of the sgRNA1 site were confirmed.These findings underscore the viability of the hairy root transformation system in assessing the editing efficiency of sgRNA targets in diverse Citrullus species.
文摘The artemisinin accumulation in the hairy root cultures of Artemisia annua L. was enhanced via a treatment of three fungal elicitors separately ( Verticillium dahliae Kleb., Rhizopus stolonifer (Ehrenb. ex Fr.) Vuill and Colletotrichum dematium (Pers.) Grove). Among these three elicitors, V. dahliae had the highest inducing efficiency, but none of them manifests any noticeable effects on the cell growth of the hairy root cultures. The artemisinin content of the hairy root cultures treated with V. dahliae elicitor was 1.12 mg/g DW, which was 45% higher than the control (0.77 mg/g DW). The results showed that elicitation was dependent on the elicitor concentration, the incubation period and the physiological stage at which the hairy root cultures were treated. In addition, the authors found that for V. dahliae , the optimum concentration was 0.4 mg carbohydrate per millilitre medium, the strongest response of A. annua hairy root cultures to the elicitation was at the late exponential growth stage, and the highest artemisinin content of the hairy root cultures was on the 4th day post treatment.
文摘The biotransformation of artemisinin by hairy root cultures ofRheum palmatum L. was investigated for the first time. The main product, deoxyartemisinin, was isolated and characterized on the basis of its spectral data.
文摘The Hairy Ape is a representative play of Eugene O'Neill that distinguishes collective trauma of mechanization and materialism in the 20 th century of America. Till now, there have not been any thesis discussing The Hairy Ape from the perspective of trauma theory. And the studies of other works on trauma theory is also not many. Based on trauma theory, this thesis makes a decoding in The Hairy Ape.
基金supported by the National Key Research and Development Program of China (2016YFD0101005)the Agricultural Science and Technology Program for Innovation Team on Identification and excavation of Elite Crop Germplasm, Chinese Academy of Agricultural Sciences
文摘The NAC （NAM, ATAF1/2 and CUC2） transcription factor family plays a key role in plant development and responses to abiotic stress. GmNAC15 （Glyma 15g40510.1）, a member of the NAC transcription factor family in soybean, was functionally characterized, especially with regard to its role in salt tolerance. In the present study, qRT-PCR （quantitative reverse transcription PCR） analysis indicated that GmNAC15 was induced by salt, drought, low temperature stress, and ABA treatment in roots and leaves. GmNAC15 overexpression in soybean （Glycine max） hairy roots enhanced salt tolerance. Transgenic hairy roots improved the survival of wild leaves; however, overexpression of GmNAC15 in hairy root couldn＇t influnce the expression level of GmNAC15 in leaf. GmNAC15 regulates the expression levels of genes responsive to salt stress. Altogether, these results provide experimental evidence of the positive effect of GmNAC15 on salt tolerance in soybean and the potential application of genetic manipulation to enhance the salt tolerance of important crops.
文摘Black hairy tongue (BHT) is a benign medical condition characterized by elongated filiform lingual papillae with typical carpet-like appearance of the dorsum of the tongue. Its prevalence varies geographically, typically ranging from 0.6% to 11.3%. Known predisposing factors include smoking, excessive coffee/black tea consumption, poor oral hygiene, trigeminal neuralgia, general debilitation, xerostomia, and medication use. Clinical presentation varies but is typically asymptomatic, although aesthetic concerns are common. Differential diagnosis includes pseudo-BHT, acanthosis nigricans, oral hairy leukoplakia, pigmented fungiform papillae of the tongue, and congenital melanocytic/melanotic nevi/macules. Clinical diagnosis relies on visual observation, detailed history taking, and occasionally microscopic evaluation. Treatment involves identification and discontinuation of the offending agent, modifications of chronic predisposing factors, patient’s re-assurance to the benign nature of the condition, and maintenance of adequate oral hygiene with gentle debridement to promote desquamation. Complications of BHT (burning mouth syndrome, halitosis, nausea, gagging, dysgeusia) typically respond to therapy. Prognosis is excellent with treatment of underlying medical conditions. BHT remains an important medical condition which may result in additional burden on the patient and health care system and requires appropriate prevention, recognition and treatment.
基金Item supported by China national"863"high-tech program (2002AA212191)China ministry of educa-tion and science and technology commission of Shanghai(04XD14011)
文摘Catharanthus roseus contains important anti-tumor terpenoid indole alkaloids (TIAs) such as vinblastine and vincristine. Cytochrome P450 enzyme geraniol 10-hydroxylase (G10H) is a putative rate-limiting enzyme involved in the TIAs biosynthetic pathway in C. roseus. In this study the g10h gene driven by the cauliflower mosaic virus 35S (CaMV 35S) promoter was introduced into C. roseus through Agrobacterium-mediated transformation. The integration and overexpression of the target gene (g10h) in hairy root lines were confirmed by polymerase chain reaction and RT-QPCR analysis respectively. Overexpression of g10h in transgenic hairy root lines significantly enhanced the accumulations of monomeric alkaloid ajmalicine and dimeric alkaloids, vincristine and vinblastine. Total TIAs production in hairy roots reached (9.51) mg/g DW, over 3-fold higher than that in the untransformed root lines. This is the first report that engineering of g10h into TIAs-producing plant species results in significant enhancement of TIAs accumulation in cultured hairy roots. This study demonstrates that the putative rate-limiting step catalyzed by G10H is indeed the real rate-limiting step involved in the TIAs biosynthetic pathway in C. roseus, which is one of the key targets for promoting TIAs production by genetic engineering.
基金Supported by Grant from Universitas Airlangga,Surabaya,Indonesia with Grant No.8714/UN3/KR/2013
文摘Objective:To increase biomass and saponin production in hairy root culture of Talinum paniculatum Gaertn.(T.paniculatum)in balloon-type bubble bioreactor(BTBB).Methods:Hairy roots which were collected from leaf explants of T.paniculatum were infected by Agrobacterium rhizogenes strain LB510.The hairy roots were cultivated at400 m L Murashige and Skoog liquid medium without growth regulator(MS0)in1 000 m L BTBB.Each BTBB had 2 g hairy roots as initial inoculum and these cultures were treated with various concentrations of sucrose(3%,4%,5%,6%w/v)and potassium nitrate(0.5,1.0,1.5 and 2.0 strength of MS medium).Cultures were maintained for 14days.Fresh and dry weights of hairy roots at the end of culture were investigated.Results:Various concentrations of sucrose influenced the biomass accumulation of hairy roots.Maximum biomass was reached by MS medium supplemented with 6%sucrose and it was approximately threefold higher than control.Culture supplemented with potassium nitrate at 2.0 strength of MS0 could increase biomass accumulation of hairy roots until 0.14 g dry weight and it was almost threefold higher than control.However,the maximum saponin content was obtained by MS medium supplemented with 5%sucrose and 2.0 strength potassium nitrate of MS.Conclusions:Based on this research,those conditions can be used to produce biomass and saponin of hairy root of T.paniculatum in the large scale.
基金supported by the funding from the Creative Research Groups of Heilongjiang Province of China(JC2016004)the National Key R&D Program of China(2016YFD0100201-21)+1 种基金the Project of Outstanding Academic Leaders in Harbin,China(2015RQXXJ018)the China Agriculture Collaborative Creation Research System of Miscellaneous Grain Crops
文摘Glycogen synthase kinase 3(GSK3)is a kind of sedne/threonine kinase widely found in eukaryotes.Many plant GSK3 kinases play important roles in regulating stress responses.This study investigated BRASSINOSTEROID-INSENSITIVE 2(GmBIN2)gene,a member of the GSK3 protein kinase family in soybean and an orthologue of Arabidopsis BIN2/AtSK21.GmBIN2 expression was increased by salt and drought stresses,but was not significantly affected by the ABA treatment.To examine the function of GrnBIN2,transgenic Arabidopsis and transgenic soybean hairy roots were generated.Overexpression of GmBIN2 in Arabidopsis resulted in increased germination rate and root length compared with wild-type plants under salt and mannitol treatments.Overexpression of GmBIN2 increased cellular Ca2~content and reduced Na~content,enhancing salt tolerance in transgenic Arabidopsis plants.In the soybean hairy root assay,overexpression of GmBIN2 in transgenic roots also showed significantly higher relative root growth rate than the control when subjected to salt and mannitol treatments.Measurement of physiological indicators,including proline content,superoxide dismutase(SOD)activity,and relative electrical conductivity,supported this conclusion.Furthermore,we also found that GmBIN2 could up-regulate the expression of some stress-related genes in transgenic Arabidopsis and soybean hairy roots.Overall,these results indicated that GmBIN2 improved tolerance to salt and drought in transgenic Arabidopsis and soybean hairy roots.
基金supported by the Major Science and Technology Projects of China (2016ZX08010-004)the Ministry of Science and Technology of China (2016YFD0100504)the CAAS (Chinese Academy of Agriculture Sciences) Innovation Project
文摘Soybean is one of the world's most important oil and protein crops. Efficient transformation is a key factor for the improvement of soybean by genetic modification. We describe an optimized protocol for the Agrobacterium rhizogenes-mediated transformation of soybean and the induction of hairy root development in vitro. Cotyledons with 0.5-cm hypocotyls were cut from 5-day-old seedlings and used as explants. After infection and co-cultivation,hairy roots were produced in induction culture medium after 10–12 days. Using this method, 90%–99% of the infected explants of five different cultivars produced hairy roots within one month. Observations using reporter constructs showed that 30%–60% of the hairy roots induced were transformed. Based on high transformation efficiency and short transformation period, this method represents an efficient and rapid platform for study of soybean gene function.
文摘The glycosylation of hydroxylcoumarin was investigated by using suspension cultures of hairy roots of Polygonum multiflorum. Two new coumarin glucosides （3 and 4） were biosysthesized by regioselectively glycosylation of corresponding substrates （1 and 2） in the system. The structures of two products were identified as 7-hydroxy-4-methylcoumarin 5-O-β-D-glucopyranoside and 7- hydroxy-3,4-dimethylcoumarin 5-O-β-D-glucopyranoside on the ground of chemical and spectroscopic methods, respectively. 2007 Rong Min Yu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
文摘The legume forage Alhagi pseudoalhagi was transformed by the Agrobacterium rhizogenes strain A4 using cotyledon and hypocotyl segments as infection materials. Regenerated plants were achieved from sterile calli derived from hairy roots, which occurred at or near the infection sites. The regenerated plants from hairy root were characterized by normal leaf morphology and stem growth but a shallow and more extensive root system than normal plants. Opine synthesis, PCR and Southern blot confirmed that T- DNA had been integrated into the A. pseudoalhagi genome. Acetosyringone (AS) was found to be vital for successful transformation of A. pseudoalhagi.
文摘The effects of various rare earth elements on growth and isoflavonoid production in hairy root cultures of Pueraria lobata (Willd.) Ohwi (strain TR2) cultured in 500 ml flasks were studied. After 32 days of culture, the biomass of hairy roots increase 15 times and reach 3.2 g dry weight. Hairy root growth was inhibited by Y_2O_3, NaSeO_3 and Sm^(3+) because of brown formation. But significant promoting effect on root growth due to callus formation was observed in La^(3+) treatments. It is the most noteworthy that the production of total isoflavonoids and puerarin was enhanced greatly by La^(3+) treatment. A major portion of increased total isoflavonoids and puerarin was released into medium in La^(3+) treatment while the hairy root viabilities were preserved. Some specific secondary metabolite release processes could be induced by La^(3+) and their possible mechanism is discussed.
基金supported by a grant from the Distinguished Young Scholars in Universities in Liaoning Province Development Plan, No. LJQ2011090
文摘Muscle-derived stem cells were isolated from the skeletal muscle of Sprague-Dawley neonatal rats aged 3 days old. Cells at passage 5 were incubated in Dulbecco＇s modified Eagle＇s medium supplemented with 10% （v/v） fetal bovine serum, 20 IJg/L nerve growth factor, 20 pg/L basic fibroblast growth factor and 1% （v/v） penicillin for 6 days. Cells presented with long processes, similar to nerve cells. Connections were formed between cell processes. Immunocytochemical staining with neuron specific enolase verified that cells differentiated into neuron-like cells. Immunofluorescence cytochemistry and western blot results revealed that the expression of protein hairy enhancer of split-1 was significantly reduced. These results indicate that low expression of protein hairy enhancer of split-1 participates in the differentiation of muscle-derived stem cells into neuron-like cells.
文摘We present some novel rotating hairy black hole metric in （2 ＋ 1） dimensions, which is an exact solution to the field equations of the Einstein-scalar-AdS theory with a non-minimal coupling. The scalar potential is determined by the metric ansatz and consistency of the field equations and cannot be prescribed arbitrarily. In the simplified, critical ease, the scalar potential contains two independent constant parameters, which are respectively related to the mass and angular momentum of the black hole in a particular way. As long as the angular momentum does not vanish, the metric can have zero, one or two horizons. The case with no horizon is physically uninteresting because of the curvature singularity lying at the origin. We identify the necessary conditions for at least one horizon to be present in the solution, which imposes some bound on the mass-angular momentum ratio. For some particular choice of pararneters our solution degenerates into some previously known black hole solutions.
文摘Tetrahydrocannabinol (THC) derivatives are used clinically as analgesic, anti-inflammatory, appetite stimulant, anti-emetic and anti-tumor cannabinoids. THC and its related compounds are at present obtained by extraction from intact Cannabis plants or chemical synthesis, but plant cell cultures may be an alternative source of production. In the present study, hairy root cultures of C. sativa (Cannabaceae) were induced by incubation of aseptically grown callus culture with solid B5 medium supplemented with 4 mg/l naphthaleneacetic acid in darkness at 25°C. Hairy root growth profiles in shake flask, increased periodically during 35 days of growth cycle. The cannabinoid contents produced in minor levels and remained below 2.0 μg/g dry weight. The contents of can-nabinoid were analyzed by liquid chromatography and confirmed by mass spectrometry.
