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1.
The green microalga Chlorella zofingiensis can produce the ketocarotenoid astaxanthin under heterotrophic culture conditions. Here we report the growth-associated biosynthesis
of astaxanthin in this biotechnologically important alga. With glucose as sole carbon and energy source, C. zofinginesis grew fast in the dark with rapid exhaustion of nitrogen and carbon sources from media, leading to a high specific growth
rate (0.034 h−1). Cultures started at a cell concentration of about 3.4 × 109 cells l−1 reached, after 6 days, standing biomass values of 1.6 × 1011 cells or 8.5 g dry weight l−1. Surprisingly, the biosynthesis of astaxanthin was found to start at early exponential phase, independent of cessation of
cell division. A general trend was observed that the culture conditions benefiting cell growth also benefited astaxanthin
accumulation, indicating that astaxanthin was a growth-associated product in this alga. The maximum cell dry biomass and astaxanthin
yield were 11.75 g l−1 and 11.14 mg l−1 (about 1 mg g−1), simultaneously obtained in the fed-batch culture with a combined glucose–nitrate mixture addition, which were the highest
ever reported in dark-heterotrophic algal cultures. The possible reasons why dark-heterotrophic C. zofingiensis could produce astaxanthin during the course of cell growth were discussed. 相似文献
2.
Effects of nitrogen sources on cell growth and lipid accumulation of green alga <Emphasis Type="Italic">Neochloris oleoabundans</Emphasis> 总被引:8,自引:0,他引:8
Microalgal lipids are the oils of future for sustainable biodiesel production. However, relatively high production costs due
to low lipid productivity have been one of the major obstacles impeding their commercial production. We studied the effects
of nitrogen sources and their concentrations on cell growth and lipid accumulation of Neochloris oleoabundans, one of the most promising oil-rich microalgal species. While the highest lipid cell content of 0.40 g/g was obtained at
the lowest sodium nitrate concentration (3 mM), a remarkable lipid productivity of 0.133 g l−1 day−1 was achieved at 5 mM with a lipid cell content of 0.34 g/g and a biomass productivity of 0.40 g l−1 day−1. The highest biomass productivity was obtained at 10 mM sodium nitrate, with a biomass concentration of 3.2 g/l and a biomass
productivity of 0.63 g l−1 day−1. It was observed that cell growth continued after the exhaustion of external nitrogen pool, hypothetically supported by the
consumption of intracellular nitrogen pools such as chlorophyll molecules. The relationship among nitrate depletion, cell
growth, lipid cell content, and cell chlorophyll content are discussed. 相似文献
3.
Kim YS Lee JH Kim NH Yeom SJ Kim SW Oh DK 《Applied microbiology and biotechnology》2011,90(2):489-497
In the fed-batch culture of glycerol using a metabolically engineered strain of Escherichia coli, supplementation with glucose as an auxiliary carbon source increased lycopene production due to a significant increase in
cell mass, despite a reduction in specific lycopene content. l-Arabinose supplementation increased lycopene production due to increases in cell mass and specific lycopene content. Supplementation
with both glucose and l-arabinose increased lycopene production significantly due to the synergistic effect of the two sugars. Cell growth by the
consumption of carbon sources was related to endogenous metabolism in the host E. coli. Supplementation with l-arabinose stimulated only the mevalonate pathway for lycopene biosynthesis and supplementation with both glucose and l-arabinose stimulated synergistically only the mevalonate pathway. In the fed-batch culture of glycerol with 10 g l−1 glucose and 7.5 g l−1
l-arabinose, the cell mass, lycopene concentration, specific lycopene content, and lycopene productivity after 34 h were 42 g l−1, 1,350 mg l−1, 32 mg g cells−1, and 40 mg l−1 h−1, respectively. These values were 3.9-, 7.1-, 1.9-, and 11.7-fold higher than those without the auxiliary carbon sources,
respectively. This is the highest reported concentration and productivity of lycopene. 相似文献
4.
Ethrel treatment enhanced isoflavonoids accumulation in cell suspension cultures of Pueraria tuberosa, a woody legume 总被引:1,自引:0,他引:1
The cell cultures of Pueraria tuberosa, a perennial leguminous lianas, were maintained in modified MS medium (KNO3 475 mg l−1, thiamine 1 mg l−1, biotin 1 mg l−1, calcium pantothenate 1 mg l−1) containing 0.1 mg l−1 2,4,5-trichloroacetic acid and 0.1 mg l−1 kinetin. Isoflavonoids (puerarin, genistin, daidzein, genistein) accumulation in cell suspension cultures was increased by
14-fold to ~12 mg l−1 after 48 h of adding 100 μM ethrel. Ethrel inhibitors (silver nitrate and silver thiosulfate) completely inhibited this effect
in the presence of ethrel and isoflavonoids were not detected in the spent medium. The increase was dose dependent and can
be explored to trigger high yield of isoflavonoids production. 相似文献
5.
Yesenia Herrera Anthony I. Okoh Laura Alvarez Norma Robledo María R. Trejo-Hernández 《World journal of microbiology & biotechnology》2008,24(1):55-60
As part of our effort at establishing microbial consortia of relevance for the bioremediation of xenobiotics polluted environments
in Mexico, we assessed the aerobic biodegradation of 2,4-dichlorophenol (2,4-DCP) by a consortium of four Bacillus species
that were isolated from a polluted soil by enrichment using a mixture of chlorophenols. The bacterial consortium effectively
biodegraded 2-chlorophenol, 3-chlorophenol and 2,4-dichlorophenol at degradation rates of between 1.7 and 6.7 μmoles l−1 h−1. In the presence of NH4Cl or KNO2 as nitrogen sources, 2,4-DCP was variously degraded. Under both conditions, cell biomass attained highest values of 350 and 450 mg l−1 respectively, while the amounts of 2,4-DCP metabolized in 21 days reached peak values of 2.1 and 2.5 mM representing between
70 and 85% degradation respectively. Chloride releases during the same period were highest at 4.7 mM and 5.3 mM in the presence
of the two nitrogen sources. The presence of free-chloride in the culture medium had a significant impact on the catabolism
of 2,4-dichlorophenol. 相似文献
6.
In vitro cultures of Berberis buxifolia were established using thidiazuron (4.5, 23 and 45 mM) or picloram (4 and 40 mM) as plant growth regulators for sustaining
growth. For producing berberine, a two-stage culture was performed. In the first step, thidiazuron or picloram were used for
biomass production followed by the production stage where benzylaminopurine (4.4 mM) was added as a plant growth regulator.
Berberine yields (102 mg g−1 DW) and in vitro shoot cultures (200 mg g−1 DW) were significantly lower than those of whole plants in the field (416 mg g−1 DW). The highest productivity (0.18 mg 1−1 day−1) was attained using picloram (either 4 on 40 mM) in the first stage for producing biomass. 相似文献
7.
A genetic transformation system has been developed for callus cells of Crataegus
aronia using Agrobacterium
tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with
5 mg l−1 Indole-3-butyric acid (IBA) and 0.5 mg l−1 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different
types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red
colored callus was obtained on MS medium supplemented with 2 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l−1 kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli
were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l−1 hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this
is the first time to report an Agrobacterium-mediated transformation system in Crataegus
aronia. 相似文献
8.
Jaya Arora Shaily Goyal Kishan Gopal Ramawat 《In vitro cellular & developmental biology. Plant》2010,46(5):430-436
This report demonstrates the elicitation effect on growth and stilbene accumulation in cell cultures of Cayratia trifolia (Vitaceae) by an extract of the angiosperm parasite Cuscuta reflexa and salicylic acid in combination with sucrose feeding. Cell cultures of C. trifolia, a tropical liana, were maintained in liquid Murashige and Skoog's basal medium containing 0.25 mg l−1 naphthalene acetic acid, 0.2 mg l−1 kinetin with 3% sucrose and 250 mg l−1 casein hydrolysate. The cells treated with Cuscuta elicitor showed increased polyphenol oxidase activity with increasing concentration of the elicitor, while total phenol content
remained almost unchanged. Enhanced yield of stilbenes (∼8-fold) was recorded in the cells treated with 200 mg l−1
Cuscuta elicitor for 7 d. Optimum accumulation of stilbenes with a non-significant decrease in cell growth as compared with control
was recorded with the addition of 3% sucrose on the seventh day of cell culture. Addition of 3% sucrose with salicylic acid
at 500 μM and Cuscuta extract at 200 mg l−1 on the seventh day enhanced total stilbene yield up to 50.1 mg l−1, which was ∼14-fold higher than in control cultures. Piceid content increased ∼200-fold in such cultures. 相似文献
9.
Sarfraz Hussain Muhammad Arshad Baby Shaharoona Muhammad Saleem Azeem Khalid 《World journal of microbiology & biotechnology》2009,25(5):853-858
The rates of biodegradation of endosulfan by P. aeruginosa were determined with different initial endosulfan concentrations (10, 50, 100, 150, 200 and 250 mg l−1) and different growth linked kinetic models were fitted at these concentrations. At 10 mg endosulfan l−1, Monod no growth model was well fitted. Monod with growth model described the biodegradation pattern at an initial concentration
of 50, 100 and 150 mg endosulfan l−1. Significant increases of P. aeruginosa MN2B14 density in broth culture during incubation further support this result. Conversely, zero order kinetic model was well
fitted into the biodegradation data if initial endosulfan concentration was ≥200 mg endosulfan l−1. The kinetics of endosulfan biodegradation by P. aeruginosa MN2B14 in liquid broth was highly dependent upon its initial concentration. The results of this study could be employed for
predicting the persistence of endosulfan in water environment containing P. aeruginosa as an endosulfan degrading bacterium. 相似文献
10.
11.
Yantree Devi Sankar-Thomas Katja Saare-Surminski Reinhard Lieberei 《Plant Cell, Tissue and Organ Culture》2008,95(2):163-173
The present study describes a protocol for plant regeneration via somatic embryogenesis in temporary immersion system (TIS)
for Camptotheca acuminata. Somatic embryos were induced by culturing hypocotyl segments from 14-day-old in vitro grown C. acuminata seedlings in TIS. Hypocotyl segments were placed in culture vessels modified with a mechanical device to support the fixation
of explants. Cultures were maintained under a 16 h photoperiod with a light intensity of 60 μmol m−2 s−1 PPF at 25 ± 1°C. After 16 weeks of incubation embryogenic calli were formed above the edge of the mechanical device in the
basal Murashige and Skoog (MS) medium containing 35 g l−1 sucrose and without hormonal supplementation. For plantlet regeneration, somatic embryos at cotyledonary stage were cultured
in three different concentrations of 6-benzylamino-purine (0.5, 1.0 and 1.5 mg l−1 BAP) and in plant growth regulator (PGR) free medium. In general, 0.5 mg l−1 BAP was found to be the most effective concentration for growth and development of Camptotheca embryos in TIS. Conversion of somatic embryos into plantlets was also successfully achieved on sterile substrates moistened
with 0.5 mg l−1 BAP. Plantlets derived from cotyledonary embryos were rooted in vitro with 0.5 mg l−1 indole-3-butyric acid (IBA) before transfer to ex vitro conditions. 相似文献
12.
Gang-Guk Choi Byung-Hyuk Kim Chi-Yong Ahn Hee-Mock Oh 《Journal of applied phycology》2011,23(6):1031-1037
The influence of nitrogen (N) deficiency on the cell growth and intracellular lipid production of the alga Botryococcus braunii UTEX 572 was investigated. Biomass concentration and lipid content of B. braunii cultivated in modified Chu-13 medium containing 0.04, 0.37, and 3.66 mM nitrate were 0.23–0.38 g L−1 and 36–63% of dry cell weight, respectively. The specific growth rate of B. braunii reached a constant of 0.185 day−1 during cultivation with an initial nitrate feed of 3.66 mM. The maximum lipid content of B. braunii was 63% with 0.04 mM nitrate. However, the maximum lipid productivity of 0.019 g L−1 day−1 was achieved with 0.37 mM nitrate. The level of oleic acid, an important component of biodiesel, was higher at 86% of the
total fatty acids under N-limited conditions (0.04 mM nitrate) compared to 69% under N-sufficient conditions (3.66 mM nitrate).
Furthermore, expression of the stearoyl-ACP desaturase gene (sad) encoding a stearoyl-ACP desaturase involved in the synthesis of oleic acid was 2.6-fold higher under N-limited conditions
than under N-sufficient conditions. 相似文献
13.
Synergistic effect of morphactin on cytokinin-induced production of isoflavonoids in cell cultures of Pueraria tuberosa (Roxb. ex. Willd.) DC 总被引:1,自引:0,他引:1
Isoflavonoids, the functional molecules of Fabaceae, are under clinical trials against cancer, osteoporosis and cardiovascular
diseases. In this study, the efficacy of different plant growth regulators was evaluated for optimizing the production of
isoflavonoids in Pueraria tuberosa. The cultures were maintained in Murashige and Skoog’s medium containing 0.1 mg l−1 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and 0.1 mg l−1 kinetin. The addition of 5.0 mg l−1 N6-(2-Isopentenyl) adenine (2iP) resulted in about ∼32-folds increase in production of isoflavonoids, while about ∼23-folds
increase was recorded in the absence of kinetin in the maintenance medium. A maximum yield of isoflavonoids (∼80 mg l−1; 82-folds increase) was obtained in cultures grown at 0.1 mg l−1 morphactin and 5.0 mg l−1 of 2iP. However, 2,4,5-T in combination with 2iP was ineffective for their production. Among different plant growth regulators
tested, maximum yields of puerarin, genistin, daidzein and genistein were 17.4, 15.9, 69.0 and 0.04 mg l−1, respectively. The study suggested that the presence of two cytokinins or 2iP with morphactin in the culture medium markedly
enhanced the production of isoflavonoids in P. tuberosa. 相似文献
14.
Meiru Li Hongqing Li Xiaoying Hu Xiaoping Pan Guojiang Wu 《Plant Cell, Tissue and Organ Culture》2011,106(3):363-371
Saussurea involucrata is a valuable traditional Chinese medicinal herb. This is the first report of a successful genetic transformation protocol
for S. involucrata using Agrobacterium tumefaciens. Leaf explants were incubated with A. tumefaciens strain EHA105 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 as a reporter gene. Following
co-cultivation, about 23.7% of the explants produced hygromycin-resistant calli on MS basal medium (Murashige and Skoog in
Physiol Plant 15: 473–497, 1962) supplemented with 1 mg l−1 benzyladenine (BA), 0.1 mg l−1 α-naphthaleneacetic acid (NAA), 0.1 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D), 20 mg l−1 hygromycin, and 500 mg l−1 cefotaxime. Shoots were regenerated following transfer of the resistant calli to shoot induction medium containing 1.5 mg l−1 BA, 0.1 mg l−1 NAA, 0.25 mg l−1 gibberellic acid (GA3), 20 mg l−1 hygromycin, and 250 mg l−1 cefotaxime, and about 67.5% of the resistant calli differentiated into shoots. Finally, 80% of the hygromycin-resistant shoots
rooted on MS media supplemented with 0.2 mg l−1 NAA, 20 mg l−1 hygromycin, and 250 mg l−1 cefotaxime. The transgenic nature of the transformants was demonstrated by detection of β-glucuronidase activity in the primary
transformants and by Southern blot hybridization analysis. About 16% of the total inoculated leaf explants produced transgenic
plants after approximately 5 months. Using this optimized transformation system, a rice ortholog of the Arabidopsis FLOWERING LOCUS T gene, Hd3a, was transferred into S. involucrata. Introduction of this gene caused an early-flowering phenotype in S. involucrata. 相似文献
15.
Herrera-Valencia VA Contreras-Pool PY López-Adrián SJ Peraza-Echeverría S Barahona-Pérez LF 《Current microbiology》2011,63(2):151-157
The aim of this study was to investigate the potential of the green microalga Chlorella saccharophila as a source of oil for biodiesel production. We evaluated for the first time, the effect of salinity and/or nitrogen depletion
(ND) on cell growth, lipid accumulation and lipid profile in this microalga. The fatty acid methyl esters (FAME) identified
for C. saccharophila in this study consisted of C-16:0, C-18:0, C-18:1 cis, and C-18:1 trans. Among these, C-18:1 (indicator of biodiesel quality)
was the main FAME found, representing approximately 76 and 80% of total FAME under normal and ND growing conditions, respectively.
Under a normal growing condition this microalga showed 154.63 mg l−1 d−1, 63.33 mg l−1 d−1, and 103.73 mg l−1 of biomass productivity, lipid productivity, and FAME yield, respectively. The higher biomass productivity (159.58 mg l−1 d−1), lipid productivity (99.33 mg l−1 d−1), and FAME yield (315.53 mg l−1) were obtained under the ND treatment. In comparison to other related studies, our results suggest that C. saccharophila can be considered as a suitable source of oil for biodiesel production. 相似文献
16.
Rhamnolipid biosurfactant production by Pseudomonas nitroreducens isolated from petroleum-contaminated soil was investigated. The effects of carbon, nitrogen and carbon to nitrogen ratio
on biosurfactant production were examined using mineral salts medium as the growth medium. The tenso-active properties (surface
activity and critical micelle concentrations of the produced biosurfactant were also evaluated. The best carbon source, nitrogen
source were glucose and sodium nitrate giving rhamnolipid yields of 5.28 and 4.38 g l−1, respectively. The maximum rhamnolipid production of 5.46 g l−1 was at C/N (glucose/sodium nitrate) of 22. The rhamnolipid biosurfactant reduced the surface tension of water from 72 to
~37 mN/m. It also has critical micelle concentration of ~28 mg l−1. Thus, the results presented in our reports show that the produced rhamnolipid can find wide applications in various bioremediation
activities such as enhanced oil recovery and petroleum degradation. 相似文献
17.
Hong-Sheng Wu Waseem Raza Dong-Yang Liu Cheng-Long Wu Ze-Shen Mao Yang-Chun Xu Qi-Rong Shen 《World journal of microbiology & biotechnology》2008,24(8):1297-1304
Watermelon production is threatened by fusarium wilt caused by Fusarium oxysporum f.sp. niveum (FON) in continuous cultivation system. Some elements, mainly allelochemicals, released from living roots or decayed plants
might be associated with the disease. The purpose of this work was to evaluate the possible impact of coumarin, one kind of
watermelon allelochemical, on FON. Furthermore, possible new mechanisms might be investigated during the ecological interactions
of plant-microbe. Results showed that coumarin strongly inhibited growth of FON leading to a decrease in its biomass, dry
weight of mycelia of FON in a liquid culture. The dry weight was decreased by 62.9% compared with control. The hyphal growth
of FON on plates was stopped at high (>400 mg l−1) concentrations of coumarin. At 320 mg l−1, sporulation and enzyme activities of FON were also severely suppressed by coumarin. The yield of conidia, and the activities
of proteinase, cellulase, and amylase were reduced by 98.9%, 79.7%, 29.8% and 15.9% respectively. However, conidial germination
and mycotoxin (MT) production of FON were greatly stimulated, being increased by 55.7% and 14.9 fold at 320 mg l−1 respectively. We conclude that coumarin acted as an allelochemical substance to inhibit growth and pathogenic enzyme activities
of FON but to stimulate mycotoxin production and conidial germination. It was suggested that coumarin acted as a signal transduction
element bridging plant and pathogen in the process of plant-microbe interactions. 相似文献
18.
Ehsan Ullah Khan Xing-Zheng Fu Ji-Hong Liu 《Plant Cell, Tissue and Organ Culture》2012,109(2):383-390
In this study, attempts were made to develop a protocol for regeneration of transgenic plants via Agrobacterium tumefaciens-mediated transformation of leaf segments from ‘Valencia’ sweet orange (Citrus sinensis L. Osbeck) using gfp (green fluorescence protein) as a vital marker. Sensitivity of the leaf segments regeneration to kanamycin was evaluated,
which showed that 50 mg l−1 was the best among the tested concentrations. In addition, factors affecting the frequency of transient gfp expression were optimized, including leaf age, Agrobacterium concentration, infection time, and co-cultivation period. Adventitious shoots regenerated on medium containing Murashige
and Tucker basal medium plus 0.1 mg l−1 α-naphthaleneacetic acid (NAA), 0.5 mg l−1 6-benzyladenine (BA) and 0.5 mg l−1 kinetin (KT). The leaf segments from 3-month-old in vitro seedlings, Agrobacterium concentration at OD600 of 0.6, 10-min immersion, and co-cultivation for 3 days yielded the highest frequency of transient gfp expression, shoots regeneration response and transformation efficiency. By applying these optimized parameters we recovered
independent transformed plants at the transformation efficiency of 23.33% on selection medium (MT salts augmented with 0.5 mg l−1 BA, 0.5 mg l−1 KT, 0.1 mg l−1 NAA, 50 mg l−1 kanamycin and 250 mg l−1 cefotaxime). Expression of gfp in the leaf segments and regenerated shoots was confirmed using fluorescence microscope. Polymerase chain reaction (PCR)
analysis using gfp and nptII gene-specific primers further confirmed the integration of the transgene in the independent transgenic plants. The transformation
methodology described here may pave the way for generating transgenic plants using leaf segments as explants. 相似文献
19.
Morinda citrifolia adventitious roots were cultured in shake flasks using Murashige and Skoog medium with different types and concentrations
of auxin and cytokinin. Root (fresh weight and dry weight) accumulation was enhanced at 5 mg l−1 indole butyric acid (IBA) and at 7 and 9 mg l−1 naphthalene acetic acid (NAA). On the other hand, 9 mg l−1 NAA decreased the anthraquinone, phenolic and flavonoid contents more severely than 9 mg l−1 IBA. When adventitious roots were treated with kinetin (0.1, 0.3 and 0.5 mg l−1) and thidiazuron (TDZ; 0.1, 0.3 and 0.5 mg l−1) in combination with 5 mg l−1 IBA, fresh weight and dry weight decreased but secondary metabolite content increased. The secondary metabolite content (including
1,1-diphenyl-2-picrylhydrazyl activity) increased more in TDZ-treated than in kinetin-treated roots. Antioxidative enzymes
such as catalase (CAT) and guaiacol peroxidase (G-POD), which play important roles in plant defense, also increased. A strong
decrease in ascorbate peroxidase activity resulted in a high accumulation of hydrogen peroxide. This indicates that adventitious
roots can grow under stress conditions with induced CAT and G-POD activities and higher accumulations of secondary metabolites.
These results suggest that 5 mg l−1 IBA supplementation is useful for growth and secondary metabolite production in adventitious roots of M. citrifolia. 相似文献
20.
Meiru Li Hongqing Li Xiaoying Hu Xiaoping Pan Guojiang Wu 《Plant Cell, Tissue and Organ Culture》2010,102(3):321-327
Zoysia tenuifolia Willd. ex Trin. is one of the most popularly cultivated turfgrass. This is the first report of successful plant regeneration
and genetic transformation protocols for Z. tenuifolia using Agrobacterium tumefaciens. Initial calli was induced from stem nodes incubated on a Murashige and Skoog (1962) (MS) medium supplemented with 2 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 1 mg l−1 6-benzyladenine (BA), with a frequency of 53%. Compact calli were selected and subcultured monthly on the fresh medium. Sixty-nine
percent of the calli could be induced to regenerate plantlets when the calli incubated on a MS medium supplemented with 0.2 mg l−1 BA under darkness. For genetic transformation, calli were incubated with A. tumefaciens strain EHA105 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, about 12% of the callus explants produced hygromycin resistant calli on
MS medium supplemented with 2 mg l−1 2,4-D, 1 mg l−1 BA, 50 mg l−1 hygromycin, 500 mg l−1 cefotaxime after 8 weeks. Shoots were regenerated following transfer of the resistant calli to shoot induction medium containing
0.2 mg l−1 BA, 50 mg l−1 hygromycin, and 250 mg l−1 cefotaxime, and about 46% of the resistant calli differentiated into shoots. Finally, all the resistant shoots were rooted
on 1/2 MS media supplemented with 50 mg l−1 hygromycin, 250 mg l−1 cefotaxime. The transgenic nature of the transformants was demonstrated by the detection of β-glucuronidase activity in the primary transformants and by PCR and Southern hybridization analysis. About 5% of the total
inoculated callus explants produced transgenic plants after approximately 5 months. The procedure described will be useful
for both, the introduction of desired genes into Z. tenuifolia and the molecular analysis of gene function. 相似文献