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1.
Alagarsamy Karthikeyan Shumugiah Karutha Pandian Manikandan Ramesh 《Plant Cell, Tissue and Organ Culture》2011,107(3):383-395
To develop salt tolerant rice, the P5CS gene of Vigna aconitifolia, encoding for proline synthesis, was introduced into the popular indica rice cultivar ADT 43. Agrobacterium tumefaciens strain LBA 4404 harboring the binary vector pCAMBIA 1301/P5CS, carrying the proline synthesis encoding gene P5CS, was co-cultivated with embryogenic callus of rice. Adding 100 μM acetosyringone to the Linsmaier and Skoog (LS) liquid and
solid co-culture medium, along with 30 mg/l hygromycin and 250 mg/l timentin, contributed to significantly higher efficiency
of transformation. Southern blot analysis of T1 independent transformants revealed that the copy number of transgene varied between one and three. When transgenic plants
were subjected to salt stress, these plants grew well in the presence of up to 200 mM NaCl, while control plants died within
10 days under these treatment conditions. These transgenic plants grew under salt stress for a period of 4 weeks, and were
capable of flowering and set seed. T1 plants segregated into 3:1 ratio suggesting Mendelian segregation pattern of inheritance of the P5CS transgene. 相似文献
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Pooja Bhatnagar-Mathur Vincent Vadez M. Jyostna Devi M. Lavanya G. Vani Kiran K. Sharma 《Molecular breeding : new strategies in plant improvement》2009,23(4):591-606
Abiotic stresses including water deficit severely limits crop yields in the semi-arid tropics. In chickpea, annual losses
of over 3.7 million tones have been estimated to be due to water deficit conditions alone. Therefore, major efforts are needed
to improve its tolerance to water deficit, and genetic engineering approaches provide an increasing hope for this possibility.
We have used transgenic technology for the introduction of an osmoregulatory gene P5CSF129A encoding the mutagenized Δ1-pyrroline-5-carboxylate synthetase (P5CS) for the overproduction of proline. A total of 49 transgenic events of chickpea were produced with the 35S:P5CSF129A gene
through Agrobacterium tumefaciens-mediated gene transfer through the use of axillary meristem explants. Eleven transgenic events that accumulated high proline
(2–6 folds) were further evaluated in greenhouse experiments based on their transpiration efficiency (TE), photosynthetic
activity, stomatal conductance, and root length under water stress. Almost all the transgenic events showed a decline in transpiration
at lower values of the fraction of transpirable soil water (dryer soil), and extracted more water than their untransformed
parents. The accumulation of proline in the selected events was more pronounced that increased significantly in the leaves
when exposed to water stress. However, the overexpression of P5CSF129A gene resulted only in a modest increase in TE, thereby indicating that the enhanced proline had little bearing on the components
of yield architecture that are significant in overcoming the negative effects of drought stress in chickpea. 相似文献
4.
Vinay Kumar Varsha Shriram P. B. Kavi Kishor Narendra Jawali M. G. Shitole 《Plant biotechnology reports》2010,4(1):37-48
Δ1-pyrroline-5-carboxylate synthetase (P5CS) is a proline biosynthetic pathway enzyme and is known for conferring enhanced salt
and drought stress in transgenics carrying this gene in a variety of plant species; however, the wild-type P5CS is subjected
to feedback control. Therefore, in the present study, we used a mutagenized version of this osmoregulatory gene-P5CSF129A, which is not subjected to feedback control, for producing transgenic indica rice plants of cultivar Karjat-3 via Agrobacterium tumefaciens. We have used two types of explants for this purpose, namely mature embryo-derived callus and shoot apices. Various parameters
for transformation were optimized including antibiotic concentration for selection, duration of cocultivation, addition of
phenolic compound, and bacterial culture density. The resultant primary transgenic plants showed more enhanced proline accumulation
than their non-transformed counterparts. This proline level was particularly enhanced in the transgenic plants of next generation
(T1) under 150 mM NaCl stress. The higher proline level shown by transgenic plants was associated with better biomass production
and growth performance under salt stress and lower extent of lipid peroxidation, indicating that overproduction of proline
may have a role in counteracting the negative effect of salt stress and higher maintenance of cellular integrity and basic
physiological processes under stress. 相似文献
5.
Proline accumulations in abiotically stressed plants is generally considered to benefit their stress tolerance. The Δ1-Pyrroline-5-carboxylate synthetase (P5CS) gene family, which encodes the rate-limiting enzyme in proline biosynthesis pathway, usually contains two duplicated genes in most plants. However, three P5CS genes including LrP5CS1, LrP5CS2 as well as a third one, LrP5CS3, were isolated from Lilium regale. LrP5CS3 is highly identical to LrP5CS1 in amino acid sequences, indicating they could come from a paralogous duplication. The phylogenetic tree suggested that the duplication of LrP5CS occurred independently after the divergence of Liliales and commelinoids. The expression of LrP5CS1 was strongly induced in leaves and roots both under drought and salinity, while that of LrP5CS3 was upregulated more moderately. LrP5CS2 stayed almost constitutive under stress. LrP5CS1 exhibited the highest activity after expressed in E. coli. Overexpression of LrP5CS genes conferred enhanced osmotic, drought and salt tolerance on transgenic Arabidopsis without negative effects in unstressed condition. Under salt stress, lines LrP5CS2 accumulated fewer proline than others, and lines LrP5CS1 grew better in root elongation. The roots of lines LrP5CS3 grew better than all others under unstressed condition and osmotic stress. Our study suggests that the three LrP5CS genes play distinct roles respectively in proline accumulation and abiotic stress tolerance. 相似文献
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Meiru Li Xiaojie Lin Hongqing Li Xiaoping Pan Guojiang Wu 《Plant Cell, Tissue and Organ Culture》2011,107(2):283-293
Overexpression of NHX genes has been previously shown to improve salt tolerance of transgenic plants. In this study, transgenic rice plants overexpressing AtNHX5 showed not only high salt tolerance, but also high drought tolerance. Measurements of ion levels indicated that Na+ and K+ contents were all higher in AtNHX5 overexpressing shoots than in wild type (WT) shoots in high saline conditions. After exposure to water deficiency and salt stress, the WT plants all died, while the AtNHX5 overexpressing rice plants had a higher survival rate, dry weight, leaf water content, and leaf chlorophyll contents, accumulated more proline, and had less membrane damage than the WT plants. In addition, seeds of both transgenic and WT plants germinated on 1/2 MS medium supplemented with 250 mM mannitol, but overexpression of AtNHX5 improved the shoot growth of the seedlings. Taken together, the results indicate that AtNHX5 gene could enhance the tolerance of rice plants to multiple environmental stresses by promoting the accumulation of more effective osmolytes (ions or proline) to counter the osmotic stress caused by abiotic factors. 相似文献
8.
V. Prabhavathi J.S. Yadav P.A. Kumar M.V. Rajam 《Molecular breeding : new strategies in plant improvement》2002,9(2):137-147
In the present work, the bacterial mannitol-1-phosphodehydrogenase(mtlD) gene was introduced into eggplant(Solanummelongena L.) by Agrobacteriumtumefaciens-mediated transformation. Several transformants weregenerated and the transgene integration was confirmed by PCR, dot blot andSouthern blot analysis. Transgenic lines of T0 and T1generations were examined for tolerance to NaCl-induced salt stress,polyethylene glycol-mediated drought and chilling stress under bothinvitro and in vivo growth conditions. Aconsiderable proportions of transgenic seeds germinated and seedlings grew wellon 200 mM salt-amended MS basal medium, whereas seeds ofuntransformed control plants failed to germinate. Further, leaf explants fromthe transgenics could grow and showed signs of shoot regeneration onsalt-amended MS regeneration medium, whereas wild type did not respond, and infact the explants showed necrosis and loss of chlorophyll after about one week.The transgenic leaves could also withstand desiccation, and transgenics couldgrow well under chilling stress, and hydroponic conditions with salt stress ascompared to wild type plants. Thus, the transgenic lines were found to betolerant against osmotic stress induced by salt, drought and chilling stress.The morphology of the transgenic plants was normal as controls, but thechlorophyll content was higher in some of the lines. These observations suggestthat mtlD gene can impart abiotic stress tolerance ineggplant. 相似文献
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Julia Tufino Silva Guerzoni Nathalia Geraldo Belintani Rosangela Maria Pinto Moreira Andrea Akemi Hoshino Douglas Silva Domingues João Carlos Bespalhok Filho Luiz Gonzaga Esteves Vieira 《Acta Physiologiae Plantarum》2014,36(9):2309-2319
High salinity interferes in sugarcane growth and development, affecting not only crop yield but also reducing sucrose concentration in culms. Sugarcane plants submitted to salt stress can accumulate compatible solutes, such as proline, which may counteract the effects of salt accumulation in the vacuole and scavenge reactive oxygen species. The objective of this study was to evaluate the response to salt stress of sugarcane plants transformed with the Vigna aconitifolia P5CS gene, which encodes ?1-pyrroline-5-carboxylate synthetase, under the control of a stress-induced promoter AIPC (ABA-inducible promoter complex). For this, 4-month-old clonally multiplied sugarcane plants from two transformation events were irrigated every 2 days with 1/10 Hoagland’s solution supplemented with 100, 150 and 200 NaCl, progressively, during 28 days. Transgenic lines showed increased transgene expression in 3.75-fold when compared with the control plants after 9 days of irrigation with saline water, which can explain the higher proline concentration found in these plants. At the end of the experiment (day 28), the transgenic lines accumulated up to 25 % higher amounts of proline when compared with non-transformed control plants. Stress response in transgenic plants was also accompanied by a reduction of malondialdehyde (MDA) derived from cellular lipid peroxidation in leaves, lower Na+ accumulation in leaves and maintenance of photochemical efficiency of PSII. Thus, proline contributed to the protection of the photosynthetic apparatus and the prevention of oxidative damage in transgenic sugarcane under salt stress. 相似文献
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Ch. Surekha K. Nirmala Kumari L. V. Aruna G. Suneetha A. Arundhati P. B. Kavi Kishor 《Plant Cell, Tissue and Organ Culture》2014,116(1):27-36
Abiotic stress is the major limiting factor of plant growth and crop yield which can be improved by osmoprotectants. Proline acts as an osmoprotectant and plays an important role in osmotic balancing, protection of sub-cellular structures, enzymes and in increasing cellular osmolarity that provide the turgor necessary for cell expansion under stress conditions. ?1-pyrroline-5-carboxylate synthetase (P5CS), a rate-limiting enzyme in proline biosynthesis which is known for conferring enhanced salt and drought stress is subjected to feedback inhibition by proline. Therefore, in the present study, we used a mutagenized version P5CSF129A of wild P5CS which is not subjected to feedback control. Efficient in vitro transformation of embryonic structures of pigeonpea (Cajanus cajan (L.) Millsp.) was obtained using Agrobacterium tumefaciens strain LBA4404 harbouring a modified binary vector pCAMBIA 1301 carrying the hptII gene for resistance to hygromycin sulphate, GUS reporter gene, encoding β-glucuronidase, and the Vigna aconitifolia P5CSF129A genes under a constitutive 35S promoter. Embryonic structures showed blue color when tested for GUS after first cycle of antibiotic selection. Integration of T-DNA into nuclear genome of transformed plants and its sexual transmission to the progeny of the transgenic plants are confirmed by PCR amplification of 340 bp hptII, 800 bp P5CSF129A fragments and Southern blot hybridization analysis. The resultant primary transgenic plants showed more proline accumulation than their non-transformed plants. Levels of proline were also elevated in T1 transgenic plants when grown in the presence of 200 mM NaCl. In addition to their enhanced growth performance, more chlorophyll and relative water content under high salinity, these plants also had lower levels of lipid peroxidation. This suggests that overproduction of proline might play an important role against salt shock and cellular integrity. 相似文献
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Vinayak H. Lokhande Ashish K. Srivastava Sudhakar Srivastava Tukaram D. Nikam Penna Suprasanna 《Plant Growth Regulation》2011,65(2):287-298
The present work addresses the importance of antioxidant, redox and energetic parameters in regulating salt-tolerance in Sesuvium portulacastrum. Experiments were conducted on 45 days old plants subjected to 250 and 1,000 mM NaCl stress for 2–8 days. Plants showed no
significant change in growth parameters (shoot length, dry weight, and water content) at 250 mM NaCl as compared to control.
However, growth of plants was significantly affected at 1,000 mM NaCl. The differential growth behaviour could be attributed
to a greater decline in the energetic parameters (in terms of ratios of NADP/NADPH and ATP/ADP) at 1,000 mM NaCl than at 250 mM
NaCl. The osmotic stress imposed to plants at 250 mM NaCl was presumably balanced by the accumulation of sodium ions (Na+), an energetically favorable process, and did not require an increased synthesis of proline. In contrast, to counter osmotic
stress at 1,000 mM NaCl, plants accumulated Na+ as well as proline and were, therefore, energetically stressed. Further, the response of enzymatic and molecular antioxidants
at 1,000 mM was either close to or even lower than that at 250 mM, which resulted in oxidative damage at 1,000 mM, particularly
on longer durations. In conclusion, it is suggested that altered redox and energetic status of the plants could play a key
role in mediating the tolerance of Sesuvium under salinity stress. 相似文献
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A. Yamchi F. Rastgar Jazii A. Mousavi A. A. Karkhane Renu 《Journal of plant biochemistry and biotechnology.》2007,16(1):9-15
The entire coding sequence of the bi-functional enzyme, Δ1-Pyrroline-5-carboxylate synthetase (P5CS) from Arabidopsis thaliana was reverse-transcribed, amplified and expressed under the control of CaMV 35S promoter in transgenic tobacco plants. Several lines were established and tested for the expression of P5CS. Drought and salinity were applied as osmotic stresses and proline content of the transformed plants was compared with that of non-transformed controls. Results indicate that transgenic lines express higher levels of proline and show enhanced resistance to the applied osmotic stress as compared to the non-transgenic plants. 相似文献
16.
Using an Agrobacterium-mediated transformation method based on wounding cultured immature seeds with carborundum (600 mesh) in liquid, auxin-regulated tobacco glutathione S -transferase (GST) (NT107) constructs were used to transform Dianthus superbusL. A 663 bp DNA band was found in the transgenic plant genome by PCR analysis using NT107-1 and NT107-2 primers, and a Southern blot analysis showed that the DIG-labelled GST gene was hybridized to the expected amplified genomic DNA fragment from transgenic D. superbus. An overexpression of NT107 led to a twofold increase in GST-specific activity compared to the non-transgenic control plants, and the GST overexpression plants showed an enhanced acclimatization in the soil. To investigate whether an increased expression of GST could affect the resistance of photosynthesis to environmental stress, these plants were subjected to drought and various light intensities from 100 to 3000 mol m–2s–1. Copper accumulation and the translocation rate were also analysed in the transgenic lines, and the GST overexpression plants were found to synthesize phytochelatin (PC), which functions by sequestering and detoxifying excess copper ions.These two authors contributed equally to this work 相似文献
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Transgenic sweetpotato (cv. Lizixiang) plants exhibiting enhanced salt tolerance were developed using LOW OSMOTIC STRESS 5 (LOS5) with Agrobacterium tumefaciens-mediated transformation. A. tumefaciens strain EHA105 harbors the pCAMBIA1300 binary vector with the LOS5 and hygromycin phosphotransferase II (hptII) genes. Selection culture was conducted using 25 mg l−1 hygromycin. A total of 26 plants were produced from the inoculated 200 cell aggregates of Lizixiang via somatic embryogenesis.
PCR analysis showed that 23 of the 26 regenerated plants were transgenic plants. All of the transgenic plants exhibited higher
salt tolerance compared to the untransformed control plants by in vitro assay for salt tolerance with 86 mM NaCl. When plants
were exposed to 86 mM NaCl, 16 transgenic plants had significantly higher levels of superoxide dismutase (SOD), proline, and
abscisic acid (ABA) and significantly lower malonaldehyde (MDA) contents than those in untransformed control plants. Salt
tolerance of these 16 plants was further evaluated with Hoagland solution containing 86 mM NaCl in a greenhouse. Four of the
sixteen had significantly better growth and rooting ability than the remaining 12 plants and control plants. Stable integration
of the LOS5 gene into the genome of the 4 salt-tolerant transgenic plants was confirmed by Southern blot analysis, and the copy number
of integrated LOS5 gene ranged from 1 to 3. High level of LOS5 gene expression in the 4 salt-tolerant transgenic plants was demonstrated by real-time quantitative PCR analysis. This study
provides an important approach for improving salt tolerance of sweetpotato. 相似文献
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Andreia Carina Turchetto-Zolet Marcia Margis-Pinheiro Rogerio Margis 《Molecular genetics and genomics : MGG》2009,281(1):87-97
Many plants synthesize and accumulate proline in response to osmotic stress conditions. A central enzyme in the proline biosynthesis
is the bifunctional enzyme Δ1-pyrroline-5-carboxylate synthase (P5CS) that includes two functional catalytic domains: the γ-glutamyl kinase and the glutamic-γ-semialdehyde
dehydrogenase. This enzyme catalyzes the first two steps of the proline biosynthetic pathway and plays a central role in the
regulation of this process in plants. To determine the evolutionary events that occurred in P5CS genes, partial sequences from four Neotropical trees were cloned and compared to those of other plant taxa. Molecular phylogenetic
analysis indicated that P5CS duplication events have occurred several times following the emergence of flowering plants and at different frequencies throughout
the evolution of monocots and dicots. Despite the high number of conserved residues in plant P5CS sequences, positive selection was observed at different regions of P5CS paralogous genes and also when dicots and monocots were contrasted. 相似文献
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S. S. Ibragimova Ya. S. Kolodyazhnaya S. V. Gerasimova A. V. Kochetov 《Russian Journal of Plant Physiology》2012,59(1):88-96
The role of gene of proline dehydrogenase (PDH) in the maintenance of stress tolerance was investigated using the model transgenic
plants of tobacco (Nicotiana tabacum L.) carrying an antisense suppressor of PDH gene (a fragment of Arabidopsis PDH gene under the control of cauliflower mosaic virus 35S promoter in antisense orientation) and notable for a low activity
of PDH and elevated content of proline. The progeny of transgenic plants belonging to the 5th generation (T5) with partially suppressed PDH activity was more resistant to various types of stress as compared with the control plants of tobacco, cv. Petit Havana SR-1
(SR1). The seedlings of transgenic lines cultured in Petri dishes on agar media supplemented with stress agents were resistant
to high NaCl concentrations (200–300 mM) and water deficit simulated by an increased agar content in the medium (14 g/l) as
compared to the control seedlings of cv. SR1. Juvenile plants of transgenic lines grown in pots filled with a mixture of vermiculite
and perlite also manifested the higher resistance to water deficit and low temperatures (2°C and −2°C) than the control plants.
Thus, the partial PDH suppression correlated with an increase in nonspecific resistance to different types of abiotic stress: salinity, water deficit,
and low temperatures. Such transgenic lines of tobacco are promising genetic models for thorough investigation of molecular
mechanisms of stress resistance in plants. 相似文献