The floral-dip transformation, the simplest technique, is no requirement of tissue culture procedure, and can directly transfer the interest gene into plant reproductive cells. It has been successfully applied to vari...The floral-dip transformation, the simplest technique, is no requirement of tissue culture procedure, and can directly transfer the interest gene into plant reproductive cells. It has been successfully applied to various plant species. In this study, the optimal conditions of a floral-dip method for production of transgenic rice variety RD41 were explored. The simple and effective inoculation medium was composed of Murashige and Skoog(MS) medium, 5% sucrose, 44 nmol/L benzylaminopurine, and 0.075% surfactant Tween-20 with pH 5.7. The transformation efficiencies of Agrobacterium tumefaciens strains AGL1 and EHA105 were compared with the Agrobacterium density at OD_(600) = 0.8–1.0 and the co-cultivation at 25 ℃ for 48 h. A. tumefaciens strain EHA105 gave slightly higher transformation efficiency than AGL1, with statistically non-significant difference. The floral-drop transformation using the optimal floral-dip conditions showed higher transformation efficiency than the floral-dip method, but the dropped flowers turned brown and died within 2 d. Production of transgenic rice variety RD41 by the floral-dip method was achieved using A. tumefaciens strain EHA105 with the optimal conditions. Screening for the gus A gene by PCR using the gus A specific primers in the T_0 lines, there were 4 transgenic lines from 286 T_0 lines(1.4% transformation efficiency). However, histochemical glucuronidase(GUS) assay demonstrated that only three of four transgenic lines exhibited gus A expression. These results indicated that floral-dip transformation is a potential tool for production of the transgenic rice, which can be used for molecular breeding via genetic engineering in the future.展开更多
Cotton plants are recalcitrant with regards to transformation and induced regeneration.In the present study,5-enolpyruvylshikimate-3-phosphate（EPSPS）,a glyphosate resistant gene from the bacterium Agrobacterium sp.s...Cotton plants are recalcitrant with regards to transformation and induced regeneration.In the present study,5-enolpyruvylshikimate-3-phosphate（EPSPS）,a glyphosate resistant gene from the bacterium Agrobacterium sp.strain CP4,was introduced into an elite Bt transgenic cotton cultivar with a modified technique involving in planta Agrobacteriummediated transformation of shoot apex.Primary transformants were initially screened using a 0.26%glyphosate spray and subsequently by PCR analysis.Five out of 4 000 transformants from T_1 seeds were obtained resulting in an in planta transformation rate of 0.125%.Four homozygous lines were produced by continuous self-fertilization and both PCR-based selection and glyphosate resistance.Transgene insertion was analyzed by Southern blot analysis.Gene transcription and protein expression levels in the transgenic cotton lines were further investigated by RT-PCR,Western blot,and ELISA methods.Transgenic T_3 plants were resistant to as much as 0.4% of glyphosate treatments in field trials.Our results indicate that the cotton shoot apex transformation technique which is both tissue-culture and genotype-independent would enable the exploitation of transgene technology in different cotton cultivars.Since this method does not require sterile conditions,the use of specialized growth media or the application of plant hormones,it can be conducted under the greenhouse condition.展开更多
Soybean transformation by ovary-drip was improved by optimizing the length of the transformation pathway by cutting the styles. These modifications facilitated soybean transformation manipulation and improved transfor...Soybean transformation by ovary-drip was improved by optimizing the length of the transformation pathway by cutting the styles. These modifications facilitated soybean transformation manipulation and improved transformation reproducibility and efficiency. Using a linear minimal gus gene cassette as the foreign DNA, a maximum transformation frequency of 11% was obtained in flowers of the soybean cultivar ‘Liaodou 14’ with their styles mostly removed, whereas removal of only the stigma, partial style cutting and partial ovary cutting gave transformation frequencies of 0%, 1%, and 2%, respectively. An average transformation frequency of 8.2% was obtained when 619 flowers from three soybean cultivars (‘Liaodou 14’, ‘Liaodou 13’, and ‘Tiefeng 29’) were transformed by this optimized method. Southern blotting analysis showed that the gus reporter gene (encoding β-glucuronidase) was stably inherited with a simple pattern. Reverse transcription-polymerase chain reaction (RT-PCR) and GUS staining confirmed the expression of the gus gene in transgenic plants.展开更多
Planta Medicais one of the leading international journals in the field of medicinal plants and natural products with original research papers,letters,rapid communications,reviews,minireviews and perspectives from rese...Planta Medicais one of the leading international journals in the field of medicinal plants and natural products with original research papers,letters,rapid communications,reviews,minireviews and perspectives from researchers worldwide.Planta Medica publishes15issues per year.展开更多
Objective To discuss the application of medial planta island flaps pedicled with anterior tibial artery perforator in front of inner malleolus for repairing small wounds around ankle Methods From Jan. 2005 to Jun. 200...Objective To discuss the application of medial planta island flaps pedicled with anterior tibial artery perforator in front of inner malleolus for repairing small wounds around ankle Methods From Jan. 2005 to Jun. 2009,10 cases with small wounds around ankle展开更多
With the development of planta genetic engineering, more emphases have been laid on convenient and high efficient genetic transformation methods. And transformation without tissue culture is a prospective direction of...With the development of planta genetic engineering, more emphases have been laid on convenient and high efficient genetic transformation methods. And transformation without tissue culture is a prospective direction of it. In this paper, traditional transformation methods and the methods of non-tissue culture were summarized. With the exploration and application of Arabidopsis transformation mechanism, with the use of ion beam-mediated transfor- mation invented by Chinese scientists and the development of other transformation methods, transformation methods without tissue culture and planta genetic engineering could be improved rapidly.展开更多
Efficient genotype-independent transformation and genome editing are highly desirable for plant biotechnology research and product development efforts.We have developed a novel approach to enable fast,high-throughput,...Efficient genotype-independent transformation and genome editing are highly desirable for plant biotechnology research and product development efforts.We have developed a novel approach to enable fast,high-throughput,and genotype-flexible Agrobacterium-mediated transformation using the important crop soybean as a test system.This new method is called GiFT(genotype-independent fast transformation)and involves only a few simple steps.The method uses germinated seeds as explants,and DNA delivery is achieved through Agrobacterium infection of wounded explants as in conventional in vitro-based methods.Following infection,the wounded explants are incubated in liquid medium with a sublethal level of selection and then transplanted directly into soil.The transplanted seedlings are then selected with herbicide spray for 3 weeks.The time required from initiation to fully established healthy T0 transgenic events is about 35 days.The GiFT method requires minimal in vitro manipulation or use of tissue culture media.Because the regeneration occurs in planta,the GiFT method is highly flexible with respect to genotype,which we demonstrate via successful transformation of elite germplasms from diverse genetic backgrounds.We also show that the soybean GiFT method can be applied to both conventional binary vectors and CRISPR-Cas12a vectors for genome editing applications.Analyses of T1 progeny demonstrate that the events have a high inheritance rate and can be used for genome engineering applications.By minimizing the need for tissue culture,the novel approach described here significantly improves operational efficiency while greatly reducing personnel and supply costs.It is the first industry-scale transformation method to utilize in planta selection in a major field crop.展开更多
Es considerada sagrada y mágica por sus peculiares propiedades.LA Rosa de Jericó(Anastatica hierochuntica)es llamada oficialmente en China como juanbai,que en espanol quiere decir “hoja de ciprés arrol...Es considerada sagrada y mágica por sus peculiares propiedades.LA Rosa de Jericó(Anastatica hierochuntica)es llamada oficialmente en China como juanbai,que en espanol quiere decir “hoja de ciprés arrollada”,pues sus hojas parecen las de ese árbol y también haber sido arrolladas.展开更多
To demonstrate the expression profiling of Xanthomonas oryzae pv.oryzae(Xoo) in vitro and in planta,DNA microarrays of 371 genes potentially associated with pathogenicity and virulence were used to compare the transcr...To demonstrate the expression profiling of Xanthomonas oryzae pv.oryzae(Xoo) in vitro and in planta,DNA microarrays of 371 genes potentially associated with pathogenicity and virulence were used to compare the transcriptional level alteration of the wild-type strain PXO99A and gene deletion mutants ΔgacAxoo and ΔfleQxoo of Xoo grown in the rich medium NBY vs.hrp-inducing minimal medium XOM2 or leaf tissues of rice.Results indicated that 17 and 38 genes of PXO99A were differentially expressed in XOM2 and the leaf tissues of rice relative to NBY,respectively.Twenty-eight genes of ΔgacAxoo grown in XOM2 and 12 genes of ΔfleQxoo in NBY were differentially expressed relative to PXO99A.The identification of differentially-expressed genes,GacAxoo-and FleQxoo-regulons and novel candidate genes of Xoo strains would provide us the target genes for further functional analysis in pathogenesis of Xoo.展开更多
基金supported by the research grant (Grant No.R2556B036) from Naresuan University, Thailand
文摘The floral-dip transformation, the simplest technique, is no requirement of tissue culture procedure, and can directly transfer the interest gene into plant reproductive cells. It has been successfully applied to various plant species. In this study, the optimal conditions of a floral-dip method for production of transgenic rice variety RD41 were explored. The simple and effective inoculation medium was composed of Murashige and Skoog(MS) medium, 5% sucrose, 44 nmol/L benzylaminopurine, and 0.075% surfactant Tween-20 with pH 5.7. The transformation efficiencies of Agrobacterium tumefaciens strains AGL1 and EHA105 were compared with the Agrobacterium density at OD_(600) = 0.8–1.0 and the co-cultivation at 25 ℃ for 48 h. A. tumefaciens strain EHA105 gave slightly higher transformation efficiency than AGL1, with statistically non-significant difference. The floral-drop transformation using the optimal floral-dip conditions showed higher transformation efficiency than the floral-dip method, but the dropped flowers turned brown and died within 2 d. Production of transgenic rice variety RD41 by the floral-dip method was achieved using A. tumefaciens strain EHA105 with the optimal conditions. Screening for the gus A gene by PCR using the gus A specific primers in the T_0 lines, there were 4 transgenic lines from 286 T_0 lines(1.4% transformation efficiency). However, histochemical glucuronidase(GUS) assay demonstrated that only three of four transgenic lines exhibited gus A expression. These results indicated that floral-dip transformation is a potential tool for production of the transgenic rice, which can be used for molecular breeding via genetic engineering in the future.
基金supported by the National Biotechnology Development Plan, China (2016ZX08005-004)
文摘Cotton plants are recalcitrant with regards to transformation and induced regeneration.In the present study,5-enolpyruvylshikimate-3-phosphate（EPSPS）,a glyphosate resistant gene from the bacterium Agrobacterium sp.strain CP4,was introduced into an elite Bt transgenic cotton cultivar with a modified technique involving in planta Agrobacteriummediated transformation of shoot apex.Primary transformants were initially screened using a 0.26%glyphosate spray and subsequently by PCR analysis.Five out of 4 000 transformants from T_1 seeds were obtained resulting in an in planta transformation rate of 0.125%.Four homozygous lines were produced by continuous self-fertilization and both PCR-based selection and glyphosate resistance.Transgene insertion was analyzed by Southern blot analysis.Gene transcription and protein expression levels in the transgenic cotton lines were further investigated by RT-PCR,Western blot,and ELISA methods.Transgenic T_3 plants were resistant to as much as 0.4% of glyphosate treatments in field trials.Our results indicate that the cotton shoot apex transformation technique which is both tissue-culture and genotype-independent would enable the exploitation of transgene technology in different cotton cultivars.Since this method does not require sterile conditions,the use of specialized growth media or the application of plant hormones,it can be conducted under the greenhouse condition.
基金Project (No. JY03-B-18-02) supported by the National R & D Project of Transgenic Crops of Ministry of Science and Technology of China
文摘Soybean transformation by ovary-drip was improved by optimizing the length of the transformation pathway by cutting the styles. These modifications facilitated soybean transformation manipulation and improved transformation reproducibility and efficiency. Using a linear minimal gus gene cassette as the foreign DNA, a maximum transformation frequency of 11% was obtained in flowers of the soybean cultivar ‘Liaodou 14’ with their styles mostly removed, whereas removal of only the stigma, partial style cutting and partial ovary cutting gave transformation frequencies of 0%, 1%, and 2%, respectively. An average transformation frequency of 8.2% was obtained when 619 flowers from three soybean cultivars (‘Liaodou 14’, ‘Liaodou 13’, and ‘Tiefeng 29’) were transformed by this optimized method. Southern blotting analysis showed that the gus reporter gene (encoding β-glucuronidase) was stably inherited with a simple pattern. Reverse transcription-polymerase chain reaction (RT-PCR) and GUS staining confirmed the expression of the gus gene in transgenic plants.
文摘Planta Medicais one of the leading international journals in the field of medicinal plants and natural products with original research papers,letters,rapid communications,reviews,minireviews and perspectives from researchers worldwide.Planta Medica publishes15issues per year.
文摘Objective To discuss the application of medial planta island flaps pedicled with anterior tibial artery perforator in front of inner malleolus for repairing small wounds around ankle Methods From Jan. 2005 to Jun. 2009,10 cases with small wounds around ankle
基金Supported by "863 Programme" (2002AA327070) and Key Technologies R&D Programme (2001BA302B-03)
文摘With the development of planta genetic engineering, more emphases have been laid on convenient and high efficient genetic transformation methods. And transformation without tissue culture is a prospective direction of it. In this paper, traditional transformation methods and the methods of non-tissue culture were summarized. With the exploration and application of Arabidopsis transformation mechanism, with the use of ion beam-mediated transfor- mation invented by Chinese scientists and the development of other transformation methods, transformation methods without tissue culture and planta genetic engineering could be improved rapidly.
基金The information described in this article is covered by two Syngenta patent publications:WO20211108336 and WO2021108337.
文摘Efficient genotype-independent transformation and genome editing are highly desirable for plant biotechnology research and product development efforts.We have developed a novel approach to enable fast,high-throughput,and genotype-flexible Agrobacterium-mediated transformation using the important crop soybean as a test system.This new method is called GiFT(genotype-independent fast transformation)and involves only a few simple steps.The method uses germinated seeds as explants,and DNA delivery is achieved through Agrobacterium infection of wounded explants as in conventional in vitro-based methods.Following infection,the wounded explants are incubated in liquid medium with a sublethal level of selection and then transplanted directly into soil.The transplanted seedlings are then selected with herbicide spray for 3 weeks.The time required from initiation to fully established healthy T0 transgenic events is about 35 days.The GiFT method requires minimal in vitro manipulation or use of tissue culture media.Because the regeneration occurs in planta,the GiFT method is highly flexible with respect to genotype,which we demonstrate via successful transformation of elite germplasms from diverse genetic backgrounds.We also show that the soybean GiFT method can be applied to both conventional binary vectors and CRISPR-Cas12a vectors for genome editing applications.Analyses of T1 progeny demonstrate that the events have a high inheritance rate and can be used for genome engineering applications.By minimizing the need for tissue culture,the novel approach described here significantly improves operational efficiency while greatly reducing personnel and supply costs.It is the first industry-scale transformation method to utilize in planta selection in a major field crop.
文摘Es considerada sagrada y mágica por sus peculiares propiedades.LA Rosa de Jericó(Anastatica hierochuntica)es llamada oficialmente en China como juanbai,que en espanol quiere decir “hoja de ciprés arrollada”,pues sus hojas parecen las de ese árbol y también haber sido arrolladas.
文摘To demonstrate the expression profiling of Xanthomonas oryzae pv.oryzae(Xoo) in vitro and in planta,DNA microarrays of 371 genes potentially associated with pathogenicity and virulence were used to compare the transcriptional level alteration of the wild-type strain PXO99A and gene deletion mutants ΔgacAxoo and ΔfleQxoo of Xoo grown in the rich medium NBY vs.hrp-inducing minimal medium XOM2 or leaf tissues of rice.Results indicated that 17 and 38 genes of PXO99A were differentially expressed in XOM2 and the leaf tissues of rice relative to NBY,respectively.Twenty-eight genes of ΔgacAxoo grown in XOM2 and 12 genes of ΔfleQxoo in NBY were differentially expressed relative to PXO99A.The identification of differentially-expressed genes,GacAxoo-and FleQxoo-regulons and novel candidate genes of Xoo strains would provide us the target genes for further functional analysis in pathogenesis of Xoo.
