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Meiru Li - One of the best experts on this subject based on the ideXlab platform.
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establishment of a highly efficient agrobacterium tumefaciens mediated leaf disc transformation method for broussonetia papyrifera
Plant Cell Tissue and Organ Culture, 2008Co-Authors: Meiru Li, Hongqing Li, Huawu Jiang, Guojiang WuAbstract:Broussonetia papyrifera is well-known for its bark fibers, which are used for making paper, cloth, rope etc. This is the first report of a successful genetic transformation protocol for B. papyrifera using Agrobacterium tumefaciens. Callus was initiated at a frequency of about 100% for both leaf and petiole explants. Shoots formed on these calli with a success rate of almost 100%, with 14.08 and 8.36 shoots regenerating from leave-derived and petiole-derived callus, respectively. For genetic transformation, leaf explants of B. papyrifera were incubated with A. tumefaciens strain LBA4404 harboring the binary vector pCAMBIA 1301 which contains the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene (gus-int) as a reporter gene. Following co-cultivation, leaf explants were cultured on Murashige and Skoog (Physiol Plant 15:473, 1962) (MS) medium supplemented with 1.5 mg l−1 benzyladenine (BA) and 0.05 mg l−1 indole-3-butyric acid (IBA) (CI medium) containing 5 mg l−1 Hygromycin and 500 mg l−1 cefotaxime, in the dark. Hygromycin-resistant calli were induced from leaf explants 3 weeks thereafter. Regenerating shoots were obtained after transfer of the calli onto MS medium supplemented with 1.5 mg l−1 BA, 0.05 mg l−1 IBA, and 0.5 mg l−1 gibberellic acid (GA3) (SI medium), 5 mg l−1 Hygromycin and 250 mg l−1 cefotaxime under fluorescent light. Finally, shoots were rooted on half strength MS medium (1/2 MS) supplemented with 10 mg l−1 Hygromycin. Transgene incorporation and expression was confirmed by PCR, Southern hybridisation and histochemical GUS assay. Using this protocol, transgenic B. papyrifera plants containing desirable new genes can be obtained in approximately 3 months with a transformation frequency as high as 44%.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
Hongqing Li - One of the best experts on this subject based on the ideXlab platform.
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establishment of a highly efficient agrobacterium tumefaciens mediated leaf disc transformation method for broussonetia papyrifera
Plant Cell Tissue and Organ Culture, 2008Co-Authors: Meiru Li, Hongqing Li, Huawu Jiang, Guojiang WuAbstract:Broussonetia papyrifera is well-known for its bark fibers, which are used for making paper, cloth, rope etc. This is the first report of a successful genetic transformation protocol for B. papyrifera using Agrobacterium tumefaciens. Callus was initiated at a frequency of about 100% for both leaf and petiole explants. Shoots formed on these calli with a success rate of almost 100%, with 14.08 and 8.36 shoots regenerating from leave-derived and petiole-derived callus, respectively. For genetic transformation, leaf explants of B. papyrifera were incubated with A. tumefaciens strain LBA4404 harboring the binary vector pCAMBIA 1301 which contains the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene (gus-int) as a reporter gene. Following co-cultivation, leaf explants were cultured on Murashige and Skoog (Physiol Plant 15:473, 1962) (MS) medium supplemented with 1.5 mg l−1 benzyladenine (BA) and 0.05 mg l−1 indole-3-butyric acid (IBA) (CI medium) containing 5 mg l−1 Hygromycin and 500 mg l−1 cefotaxime, in the dark. Hygromycin-resistant calli were induced from leaf explants 3 weeks thereafter. Regenerating shoots were obtained after transfer of the calli onto MS medium supplemented with 1.5 mg l−1 BA, 0.05 mg l−1 IBA, and 0.5 mg l−1 gibberellic acid (GA3) (SI medium), 5 mg l−1 Hygromycin and 250 mg l−1 cefotaxime under fluorescent light. Finally, shoots were rooted on half strength MS medium (1/2 MS) supplemented with 10 mg l−1 Hygromycin. Transgene incorporation and expression was confirmed by PCR, Southern hybridisation and histochemical GUS assay. Using this protocol, transgenic B. papyrifera plants containing desirable new genes can be obtained in approximately 3 months with a transformation frequency as high as 44%.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
Phillip Morris - One of the best experts on this subject based on the ideXlab platform.
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agrobacterium tumefaciens mediated transformation of festuca arundinacea schreb and lolium multiflorum lam
Plant Cell Reports, 2003Co-Authors: Andy J E Bettany, Susan Dalton, E Timms, B Manderyck, M S Dhanoa, Phillip MorrisAbstract:Agrobacterium tumefaciens strain LBA4404 carrying plasmid pTOK233 encoding the Hygromycin resistance (hph) and β-glucuronidase (uidA) genes has been used to transform two agronomic grass species: tall fescue (Festuca arundinacea) and Italian ryegrass (Lolium multiflorum). Embryogenic cell suspension colonies or young embryogenic calli were co-cultured with Agrobacterium in the presence of acetosyringone. Colonies were grown under Hygromycin selection with cefotaxime and surviving colonies plated on embryogenesis media. Eight Lolium (six independent lines) and two Festuca plants (independent lines) were regenerated and established in soil. All plants were Hygromycin-resistant, but histochemical determination of GUS activity showed that only one Festuca plant and one Lolium plant expressed GUS. Three GUS-negative transgenic L. multiflorum and the two F. arundinacea plants were vernalised and allowed to flower. All three Lolium plants were male- and female-fertile, but the Festuca plants failed to produce seed. Progeny analysis of L. multiflorum showed a 24–68% inheritance of the hph and uidA genes in the three lines with no significant difference between paternal and maternal gene transmission. However, significant differences were noted between the paternal and maternal expression of Hygromycin resistance.
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transgenic plants of lolium multiflorum lolium perenne festuca arundinacea and agrostis stolonifera by silicon carbide fibre mediated transformation of cell suspension cultures
Plant Science, 1998Co-Authors: Susan Dalton, Andy J E Bettany, E Timms, Phillip MorrisAbstract:Seven Lolium multiflorum, one Lolium perenne, 12 Festuca arundinacea and six Agrostis stolonifera plants were regenerated following transformation with a Hygromycin resistance gene and Hygromycin selection, from cell suspension colonies treated with silicon–carbide whiskers. Transformation was confirmed by PCR and Southern blotting; the latter also showed that six of the L. multiflorum plants were independent transformants (insufficient molecular evidence was obtained for the seventh), nine of the 12 F. arundinacea plants were independent transformants, but that all the A. stolonifera plants were derived from a single transformation event. Most plants tested contained fewer than five integrated transgene copies. Transgene expression was confirmed by reverse transcriptase-PCR (RT-PCR). Of the one A. stolonifera and three L. multiflorum transformants regenerated after co-transformation with both the Hygromycin resistance gene and the β-glucuronidase (gusA) gene, none were found to express GUS activity. L. multiflorum regenerants from older (14–16 week old) cell suspensions showed loss of female fertility, but analysis of the progeny from three plants showed that the transgenes were being inherited as a single dominant allele with a high frequency of transmission of Hygromycin resistance.
Tu-hsin Yan - One of the best experts on this subject based on the ideXlab platform.
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Chiral pool based efficient synthesis of the aminocyclitol core and furanoside of (-)-Hygromycin A: formal total synthesis of (-)-Hygromycin A.
Organic letters, 2012Co-Authors: Yuan-kang Chang, Tu-hsin YanAbstract:A chiral pool based synthetic strategy that leads from the readily available and inexpensive C(2)-symmetric tartaric acids to the chiral O-isopropylidenebenzooxazole--a convenient precursor to the aminocyclitol core of Hygromycin A as well as the chiral γ-disilyloxybutyrolactone--a pivotal intermediate to approach to the furanoside of Hygromycin A.
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Chiral Pool Based Efficient Synthesis of the Aminocyclitol Core and Furanoside of (−)- Hygromycin A: Formal Total Synthesis of (−)-Hygromycin A
2012Co-Authors: Yuan-kang Chang, Tu-hsin YanAbstract:A chiral pool based synthetic strategy that leads from the readily available and inexpensive C2-symmetric tartaric acids to the chiral O-isopropylidenebenzooxazolea convenient precursor to the aminocyclitol core of Hygromycin A as well as the chiral γ-disilyloxybutyrolactonea pivotal intermediate to approach to the furanoside of Hygromycin A
Lei Chen - One of the best experts on this subject based on the ideXlab platform.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
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establishment of an efficient agrobacterium tumefaciens mediated leaf disc transformation of thellungiella halophila
Plant Cell Reports, 2007Co-Authors: Hongqing Li, Jie Xu, Lei Chen, Meiru LiAbstract:Thellungiella halophila is a salt-tolerant close relative of Arabidopsis, which is adopted as a halophytic model for stress tolerance research. We established an Agrobacterium tumefaciens-mediated transformation procedure for T. halophila. Leaf explants of T. halophila were incubated with A. tumefaciens strain EHA105 containing a binary vector pCAMBIA1301 with the hpt gene as a selectable marker for Hygromycin resistance and an intron-containing β-glucuronidase gene as a reporter gene. Following co-cultivation, leaf explants were cultured on selective medium containing 10 mg l−1 Hygromycin and 500 mg l−1 cefotaxime. Hygromycin-resistant calluses were induced from the leaf explants after 3 weeks. Shoot regeneration was achieved after transferring the calluses onto fresh medium of the same composition. Finally, the shoots were rooted on half strength MS basal medium supplemented with 10 mg l−1 Hygromycin. Incorporation and expression of the transgenes were confirmed by PCR, Southern blot analysis and GUS histochemical assay. Using this protocol, transgenic T. halophila plants can be obtained in approximately 2 months with a high transformation frequency of 26%.
