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1.
High polyphenol oxidase activity and low titratable acidity in browning bamboo tissue culture 总被引:1,自引:0,他引:1
Summary Tissue browning that frequently results in the early death of bamboo shoots in vitro correlated directly with polyphenol oxidase (PPO, EC 1.10.3.1) activity and inversely with titratable acidity. It was unrelated
to the level of endogenous phenols. During the course of culture, timing of PPO activity paralleled that of explant browning.
Browning was highest among shoots cultured in a medium of pH 8, which was consistent with the pH optinum of the bamboo enzyme.
The pH optimum was first determined with the crude enzyme, then verified with two purified isozymes. Stability of the bamboo
PPO was also highest at pH 10. PPO activities of the severely browning Dendrocalamus latiflorus, the moderately browning Phyllostachys nigra, and the relatively non-browning Bambusa oldhamii were inhibited strongly by ascorbic acid, cysteine, sodium diethyldithiocarbamate, and sodium sulfite. But characterization
of bamboo PPO according to enzyme inhibitors was not possible because enzyme extracts of the three species gave varied responses
to the traditional substances. Nutrient medium addenda of some PPO inhibitors, namely ascorbic acid, cysteine, kojic acid,
and thiourea, mainly enhanced browning. However, ferulic acid at 3 mM and lower concentrations reduced the number of brown shoots per culture, although not the percentage of cultures that browned.
Polyvinylpyrrolidone failed completely to suppress browning. The two purified isozymes showed different temperature optima
for PPO activity: 60°C and 65°C. The purified isozymes displayed a substrate preference for dopamine, or a cathecol oxidase
characteristics. 相似文献
2.
V. Hadži‐Tašković Šukalović S. Veljović‐Jovanović J. Dragišić Maksimović V. Maksimović Z. Pajić 《Plant biology (Stuttgart, Germany)》2010,12(3):406-413
Silk of some maize genotypes contains a high level of phenolics that undergo enzymatic oxidation to form quinones, which condense among themselves or with proteins to form brown pigments. Two phenolic oxidizing enzymes, peroxidase (POD; EC 1.11.1.7) and polyphenol oxidase (PPO; EC 1.10.3.1), from maize (Zea mays L.) silk were characterised with respect to their preferred substrate, different isoforms and specific effectors. One browning silk sample with high, and two non‐browning samples with low phenolic content were investigated. Although POD oxidizes a wide range of phenolic substrates in vitro, its activity rate was independent of silk phenolic content. PPO activity, detected with o‐diphenolic substrates, was abundant only in browning silk, and low or absent in non‐browning silk. Pollination increased POD but not PPO activity. Isoelectric‐focusing (IEF) and specific staining for POD and PPO showed a high degree of polymorphism that varied with silk origin. The IEF pattern of POD revealed a number of anionic and several cationic isoenzymes, with the most pronounced having neutral pI 7 and a basic isoform with pI 10. Detected isoforms of PPO were anionic, except for one neutral form found only in browning silk, and occupied positions different from those of POD. Different inhibitory effects of NaN3, EDTA, KCN, and L‐cysteine, as well as different impacts of a variety of cations on the oxidation of chlorogenic acid, mediated by PPO or POD, were detected. The findings are discussed in terms of a possible roles of these enzymes in defence and pollination. 相似文献
3.
Tissue browning of in vitro cultures of Scots pine: Role of peroxidase and polyphenol oxidase 总被引:7,自引:0,他引:7
Hanna Laukkanen Hely Häggman Sari Kontunen-Soppela & Anja Hohtola 《Physiologia plantarum》1999,106(3):337-343
Callus cultures from shoot tips of mature Scots pine ( Pinus sylvestris L.) were characterized by rapid browning and an inability to regenerate. The peroxidase (POD) and polyphenol oxidase (PPO) activities and relationship to browning in such cultures were compared with embryogenic and non-embryogenic cultures of Scots pine, started from immature embryos of three different pine clones. The browning in callus cultures derived from pine buds was visible approximately after 2 weeks of culture, and continued thereafter until the callus was dark brown and poorly growing. The non-embryogenic cultures induced from immature embryos showed either light yellow coloring or browning, whereas the embryogenic cultures showed browning. POD activity increased during the first 4 weeks in callus tissue initiated from pine buds, and was significantly higher than in pine buds or cultures derived from immature embryos. The ability of cultures initiated from pine buds to oxidize catechol was notably high compared with cultures initiated from immature embryos, regardless of the time of measurement. Addition of catalase revealed that both POD and PPO were able to use catechol as substrate. An antibody raised against broad bean ( Vicia faba ) chloroplast PPO was used to recognize PPO. One polypeptide with a molecular mass of 50 kDa was detected in all pine samples on SDS-PAGE and non-denaturing PAGE. Another polypeptide with a molecular mass of 70 kDa was shown exclusively in the light-yellow non-embryogenic cultures. The results suggest that especially the high POD activities in callus tissues started from mature trees cause rapid and early browning and possibly subsequent cell death. 相似文献
4.
Hai Ping Hong Hongyi Zhang Paula Olhoft Steve Hill Hunt Wiley Effie Toren Helke Hillebrand Todd Jones Ming Cheng 《In vitro cellular & developmental biology. Plant》2007,43(6):558-568
A regeneration and transformation system has been developed using organogenic calluses derived from soybean axillary nodes
as the starting explants. Leaf-node or cotyledonary-node explants were prepared from 7 to 8-d-old seedlings. Callus was induced
on medium containing either Murashige and Skoog (MS) salts or modified Finer and Nagasawa (FNL) salts and B5 vitamins with
various concentrations of benzylamino purine (BA) and thidiazuron (TDZ). The combination of BA and TDZ had a synergistic effect
on callus induction. Shoot differentiation from the callus occurred once the callus was transferred to medium containing a
low concentration of BA. Subsequently, shoots were elongated on medium containing indole-3-acetic acid (IAA), zeatin riboside,
and gibberellic acid (GA). Plant regeneration from callus occurred 90 ∼ 120 d after the callus was cultured on shoot induction
medium. Both the primary callus and the proliferated callus were used as explants for Agrobacterium-mediated transformation. The calluses were inoculated with A. tumefaciens harboring a binary vector with the bar gene as the selectable marker gene and the gusINT gene for GUS expression. Usually 60–100% of the callus showed transient GUS expression 5 d after inoculation. Infected
calluses were then selected on media amended with various concentrations of glufosinate. Transgenic soybean plants have been
regenerated and established in the greenhouse. GUS expression was exhibited in various tissues and plant organs, including
leaf, stem, and roots. Southern and T1 plant segregation analysis of transgenic events showed that transgenes were integrated into the soybean genome with a copy
number ranging from 1–5 copies. 相似文献
5.
Yi-Ping Chen 《Plant Growth Regulation》2008,55(1):73-79
Sterilized seeds of Isatis indigotica (Brassicacae) were divided into four groups based on irradiation pretreatments. These control groups (C) were non irradiated,
He–Ne laser treated seeds (L), UV-B treated seeds (B) and He–Ne laser followed by UV-B radiation treated seeds (LB). Laser
radiation was provided by He–Ne laser, UV-B radiation was provided by filtered Qin brand 30 W fluorescent sun lamps. Malondialdehyde
(MDA), proline, UV-B absorbing compounds and ascorbic acid (AsA) concentrations, as well as, the activities of superoxide
dismutase (SOD), catalase (CAT) and peroxidase (POD) were measured in the cotyledons of seedlings from all the four irradiation
treatments. The result indicate that UV-B radiation enhanced the concentration of MDA while decreasing the activities of SOD,
CAT, POD and the concentration of AsA in the seedlings compared with the controls. The concentration of MDA decreased, while
the activities of SOD, CAT, POD and the concentration of AsA increased in seedling treated with He–Ne laser and UV-B compared
to UV-B alone. The concentration of proline and UV absorbing compounds increased progressively with treatments i.e. UV-B irradiation,
He–Ne laser irradiation, and He–Ne laser irradiation followed by UV-B irradiation compared to the controls. The present data
suggest that Isatis indigotica seedlings derived from laser stimulated seeds showed improved resistance to elevated UV-B. 相似文献
6.
In vitro microtuberization provides an adequate experimental model for the physiological and metabolic studies of tuberization and
the preliminary screenings of potential potato genotypes. The effects of saline stress at 0–80 mmol concentration on in vitro tuberization of two potato cultivars were investigated in this study. With an increase in the salt concentration, the microtuberization
of potato was either delayed by 5–10 days (20 and 40 mmol NaCl) or inhibited completely (80 mmol NaCl) in addition to the
reduction in microtuber yields. The two potato genotypes studied showed different trends in total soluble sugars, sucrose
and starch contents of microtubers under NaCl stress, while glucose and fructose levels remained unchanged. The vitamin C
content in microtubers of two potato genotypes was reduced by salt stress. Salinity applied from 20 to 60 mmol progressively
increased proline and malondialdehyde (MDA) levels in microtubers of both the potato cultivars. In genotype Zihuabai, NaCl
at a low concentration (20 mmol) led to a significant increase in peroxidase (POD) and polyphenoloxiadase (PPO) activities,
while in Jingshi-2, the PPO activity decreased progressively with an increase in NaCl concentrations. Genotype Zihuabai exhibited
higher tolerance to salt stress than Jingshi-2 under in vitro conditions. These results could be used for preliminary selections of salt tolerance in potato breeding programmes. 相似文献
7.
A transgenic apple callus showing reduced polyphenol oxidase activity and lower browning potential 总被引:3,自引:0,他引:3
Murata M Nishimura M Murai N Haruta M Homma S Itoh Y 《Bioscience, biotechnology, and biochemistry》2001,65(2):383-388
Polyphenol oxidase (PPO) is responsible for enzymatic browning of apples. Apples lacking PPO activity might be useful not only for the food industry but also for studies of the metabolism of polyphenols and the function of PPO. Transgenic apple calli were prepared by using Agrobacterium tumefaciens carrying the kanamycin (KM) resistant gene and antisense PPO gene. Four KM-resistant callus lines were obtained from 356 leaf explants. Among these transgenic calli, three calli grew on the medium containing KM at the same rate as non-transgenic callus on the medium without KM. One callus line had an antisense PPO gene, in which the amount and activity of PPO were reduced to half the amount and activity in non-transgenic callus. The browning potential of this line, which was estimated by adding chlorogenic acid, was also half the browning potential of non-transgenic callus. 相似文献
8.
Shahriar Saiedian Ezzatollah Keyhani Jacqueline Keyhani 《Acta Physiologiae Plantarum》2007,29(5):463-471
Crocus sativus L., cultivated since ancient times as the source of saffron, is a triploid plant that can be propagated only via its corms
which undergo a period of dormancy. Understanding the processes taking place in the corm is essential to preserve the plant
and improve its quality. Color and taste being of prime importance in the quality of the saffron spice, knowledge on polyphenol
oxidase (PPO) activity in the plant is of particular interest given the role of the enzyme in fruit and vegetable browning
during processing and during the storage of processed food. In this paper, PPO activity was investigated for the first time
in extracts obtained from dormant C. sativus L. corms. PPO activity was detectable using l-DOPA, pyrogallol, catechol or p-cresol as substrate, each being oxidized to its corresponding o-quinone; no activity was detectable with l-tyrosine, tyramine or phenol as substrate. Two pH optima, respectively at 4.5 and 6.7, were observed with all substrates
and a third one, at 8.5, was found with l-DOPA and p-cresol. Kinetics parameters studied at pH 6.7 indicated the highest catalytic efficiency (in units mg−1 prot mM−1) with pyrogallol: 150, then catechol: 39, l-DOPA: 6.4 and p-cresol: 4.6. The enzymatic activity was inhibited by 50% in the presence of 0.22, 0.35, 0.5 and 0.7 mM kojic acid with, respectively,
catechol, pyrogallol, p-cresol and l-DOPA as substrate. When stained for PPO activity, non-denaturing gel electropherograms of extract revealed three distinct
bands, indicating the presence of multiple isoenzymes in dormant C. sativus L. corms. 相似文献
9.
Nascimento RP d'Avila-Levy CM Souza RF Branquinha MH Bon EP Pereira-Jr N Coelho RR 《Archives of microbiology》2005,184(3):194-198
Streptomyces malaysiensis AMT-3, isolated from a Brazilian cerrado soil, showed proteolytic activities detected by gelatin–sodium dodecyl sulfate-polyacrylamide
gel electrophoresis. The optimum proteinase production was obtained when using 2.5% wheat bran and 0.1% yeast extract in the
culture medium, after 5 days incubation at 30°C. The enzymatic complex degraded gelatin optimally at pH 7.0, and under these
conditions eight proteolytic bands (four serine-proteinases and four metaloproteinases), ranging from 20 to 212 kDa, were
detected on the culture supernatant filtrates. In addition, a 35-kDa proteinase was thermostable at 60°C for 120 min. These
results point out to the applicability of gelatin zymograms in the characterization of crude enzymatic complexes. According
to our results, this enzymatic complex could be used for biotechnological applications. 相似文献
10.
Mustonen AM Saarela S Nieminen P 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》2008,178(2):199-208
Arvicolinae voles are small herbivores relying on constant food availability with weak adaptations to tolerate prolonged food
deprivation. The present study performed a comparative analysis on the responses to 4–18 h of food deprivation in the common
vole (Microtus arvalis) and the tundra vole (Microtus oeconomus). Both species exhibited rapid decreases in the plasma and liver carbohydrate concentrations during phase I of fasting and
the decline in the liver glycogen level was more pronounced in the tundra vole. The plasma thyroxine concentrations of the
common vole decreased after 4 h. Lipid mobilization (phase II of fasting) was indicated by the increased plasma free fatty
acid levels after 8–18 (the common vole) or 4–18 h (the tundra vole) and by the elevated lipase activities. In the tundra
vole, the plasma ghrelin concentrations increased after 12 h possibly to stimulate appetite. Both species showed increased
liver lipid concentrations after 4 h and plasma aminotransferase and creatine kinase activities after 12–18 h of food deprivation
implying liver dysfunction and skeletal muscle damage. No signs of stimulated protein catabolism characteristic to phase III
of fasting were present during 18 h without food. 相似文献
11.
Suxia Cui Wei Wang Chenglie Zhang 《In vitro cellular & developmental biology. Plant》2002,38(4):325-329
Summary Different ecotypes of reed (Phragmites communis Trinius) provide an ideal resource for studies on plant environmental adaptations and presence of genes relating to stress
resistance. Dune reed is a drought-tolerant reed ecotype growing in the desert regions of north-west China. In this work,
in vitro culture systems of dune reed and local swamp reed (as control) were established by optimizing the culture conditions for
each of them. Bright yellow calluses were induced on a Murashige and Skoog medium containing 4.5 μM 2,4-dichlorophenoxyacetic acid (2,4-D), 5.4 μM naphthaleneacetic acid and 2.2μM benzyladenine. Benzyladenine promoted callus induction, but was not required for callus maintenance. Four types of callus
have been identified from each of the reed ecotypes. Two types of callus, i.e. type A (formed normal green shoots) and type
C (formed albino plants), were both found as embryogenic calluses. The optimal concentrations of 2,4-D to maintain embryogenic
callus were 2.3–4.5 μM for dune reed and 9.0–13.5 μM for swap reed. Plant regeneration was achieved from types A and C callus in a hormone-free medium. The embryogenic calluses
of swamp reed have been maintained for over 2 yr and still retain their strong embryogenic potential; however, those of dune
reed gradually lost their embryogenic potential after only 7 mo. of culture. Regenerated plants from the two reed ecotypes
showed, after a growth season, similar morphology and the same chromosome number (2n=8x=96, octoploid) as the wild plants. 相似文献
12.
J. Vijaya Kumar B. D. Ranjitha Kumari G. Sujatha Enrique Castaño 《Plant Cell, Tissue and Organ Culture》2008,93(1):85-96
The present study describes a system for efficient plant regeneration via organogenesis and somatic embryogenesis of safflower
(Carthamus
tinctorius L.) cv. NARI-6 in fungal culture filtrates (FCF)-treated cultures. FCF was prepared by culturing Alternaria carthami fungal mycelia in selection medium for host-specific toxin production. Cotyledon explants cultured on callus induction medium
with different levels of FCF (10–50%) produced embryogenic callus. In organogenesis, 42.2% microshoots formed directly from
embryogenic callus tissues in plant regeneration medium with 40% FCF. Isolated embryogenic callus cultured on embryo induction
medium containing 40% FCF induced 50.2% somatic embryogenesis. Embryo germination percentage was decreased from 64.5 to 28
in embryo maturation medium containing 40% FCF. However, nine plantlets from organogenesis and 24 plantlets from somatic embryogenesis
were selected as FCF-tolerant. Alternaria carthami fungal spores (5 × 105 spores/ml) sprayed on the leaves of FCF-tolerant plants showed enhanced survival rate over control plants, which plants were
more susceptible to fungal attack. The number of leaf spot lesions per leaf was decreased from 3.4 to 0.9 and their lesion
length was also reduced from 2.9 to 0.7 mm in organogenic derived FCF-tolerant plants over control. In somatic embryo derived
FCF-tolerant plants, the number of lesions was decreased from 3.1 to 0.4 and the lesion size was also reduced to 2.7–0.5 mm
when compared to the control. This study also examined antioxidant enzyme activity in FCF-tolerant plants. Catalase (CAT)
activity was slightly decreased whereas peroxidase (POD) activity was increased to a maximum of 42% (0.19 μmol min−1 mg−1 protein) from organogenesis and 47% (0.23 μmol min−1 mg−1 protein) from embryogenesis in FCF-tolerant plants. Superoxide dismutase (SOD) activity was also increased to 17% (149 U
mg−1 protein) and 19.5% (145 U mg−1 protein) in FCF-tolerant plants derived from organogenesis and somatic embryogenesis when compared with control plants. 相似文献
13.
Ottem KF Nylund A Karlsbakk E Friis-Møller A Krossøy B Knappskog D 《Archives of microbiology》2007,188(5):547-550
A Francisella strain, GM2212, previously isolated from moribund farmed Atlantic cod (Gadus morhua) in Norway, is closely related to Francisella philomiragia among Francisella spp. according to its complete 16S rDNA, 16S-23S intergenic spacer, 23S rDNA, 23S–5S intergenic spacer, 5S rDNA, FopA, lipoprotein
TUL4 (LpnA), malate dehydrogenase and hypothetical lipoprotein (LpnB) sequences. A comparison between GM2212 and the type
strain of Francisella philomiragia were performed by DNA–DNA hybridization and fatty acid analysis. The DNA–DNA hybridization showed a 70% similarity. The fatty
acid analysis showed only minor differences between the Francisella isolates. Due to the inconclusive result from the DNA–DNA hybridisation, major emphasis concerning the status of this isolate
is made on previously published molecular, phenotypic and biochemical characters. All characteristics taken together support
the establishment of GM2212 as a novel species, for which the name Francisella piscicida sp. nov. is proposed (=CNCM I-3511T = DSM 18777T = LMG registration number not yet
available). 相似文献
14.
In the present study, the procedures for induction of somatic embryogenesis (SE) in an in vitro culture of the tulip have been developed. SE was initiated on flower stem explants isolated from “Apeldoorn” bulbs during
their low-temperature treatment. Bulbs had not been chilled or had been chilled for 12 or 24 weeks at 5°C. The explants were
cultured with exogenous auxins 2,4-dichlorophenoxyacetic acid (2,4-D), 4-amino-3,5,6-trichloropicolinic acid (Picloram), α-naphthaleneacetic
acid (NAA) at 1–100 μM and cytokinins: benzyladenine (BA) and zeatin (ZEA) at 0.5–50 μM. Increase in auxin concentrations
caused an intensive enlargement of the explant parenchyma, which changed into homogenous colorless callus. On the same media,
vein bundles developed into yellowish, nodular callus. Picloram was more efficient in inducing the formation of embryogenic
nodular callus than 2,4-D, whereas the latter stimulated formation of colorless callus. The base of the lower part of the
flower stem isolated from bulbs chilled for 12 weeks proved to be the best explant for callus formation. The highest number
of somatic embryos was produced on medium with 25 μM Picloram and 0.5 μM BA. Development of adventitious roots was noticed
in the presence of 2,4-D. Globular embryos developed into torpedo stage embryos under the influence of BA (5 μM) and NAA (0.5 μM).
Morphological and anatomical data describing development of callus and somatic embryos are presented. 相似文献
15.
壳聚糖涂膜对机械伤苹果抗性生理特征的影响 总被引:1,自引:0,他引:1
为了提高苹果采后抗机械损伤能力,减少贮藏损失,以红富士苹果为材料,通过人工模拟机械伤试验,研究壳聚糖涂膜对损伤红富士苹果常温贮藏条件下果肉褐变、相关酶活性及抗性相关物质的影响,探索壳聚糖涂膜在果蔬防腐保鲜上的应用。结果显示:壳聚糖涂膜处理科显著减少红富士苹果果实机械伤口的扩张,提高机械伤果实的总酚含量,降低PPO活性,从而有效抑制机械伤苹果贮藏期间的果肉褐变的发生。同时,壳聚糖涂膜处理可提高机械伤苹果的POD和PAL活性,延缓酚类物质含量的下降,促进体内木质素的合成。研究表明,壳聚糖涂膜处理能够有效防止机械伤苹果贮藏期间的酶促褐变,减少果肉组织中有害物质的积累,促进愈伤组织的形成,从而增强了机械伤苹果的抗性,有效延缓了果实衰老。 相似文献
16.
Biochemical features of maize tissues with different capacities to regenerate plants 总被引:4,自引:0,他引:4
Metabolic profiling using GC–MS and LC–MS analyses of soluble metabolites and cell wall bound phenolic compounds from maize calluses of different morphogenic competence revealed a number of biochemical characteristics that distinguish tissues with high plant regeneration ability from tissues that cannot efficiently regenerate plants in vitro. Maize cultures of different ages from H99 (compact type I callus) and HiII (friable type II callus) were divided into two different samples: regenerable (R) and non-regenerable (NR) based on known morphologies. Tissues from both genotypes with high morphogenic potential had higher asparagine and aspartate and indole-3-butenol concentrations, decreased sugar and DIMBOA (2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one) concentrations, low levels of 4-aminobutyric acid (GABA) and chlorogenic acid and lower levels of feruloyl- and sinapoyl glucosides compared to NR tissues. The ether bound cell wall phenolics of tissues with high regeneration potential had higher levels of the predominant G (guaiacyl) units and lower levels of H (p-hydroxyphenyl) and S (syringyl) units and higher ferulic acid/coumaric acid and ferulic acid/diferulic acid ratios. The same trends were found with the ester-bound phenolics of HiII, however, there were only small differences between the H99 R and NR tissues. Concentrations of the major sugars, organic acids, amino acids and soluble aromatic compounds tended to increase as the time after culture initiation increased. The results show that there are differences in general metabolism, phenolic secondary compounds and cell wall composition between R and NR cell types. 相似文献
17.
Lipid and docosahexaenoic acid (DHA) accumulation into Schizochytrium G13/2S was studied under batch and continuous culture. Different glucose and glutamate concentrations were supplemented in
a defined medium. During batch cultivation, lipid accumulation, 35% total fatty acids (TFA) occurred at the arithmetic growth
phase but ceased when cell growth stopped. When continuous culture was performed under different glutamate concentrations,
nitrogen-growth-limiting conditions induced the accumulation of 30–28% TFA in Schizochytrium. As the dilution rate decreased from 0.08 to 0.02 h−1, both cell dry weight and TFA content of the cell increased. Under a constant dilution rate of 0.04 h−1, carbon-limiting conditions decreased the TFA to 22%. Fatty acid profile was not affected by the different nutrient concentrations
provided during continuous culture. Consequently, lipid accumulation can be induced through the carbon and nitrogen source
concentration in the medium to maximise the TFA and subsequently DHA productivity by this microorganism. 相似文献
18.
Mature zygotic embryos of recalcitrant Christmas tree species eastern white pine (Pinus strobus L.) were used as explants for Agrobacterium tumefaciens strain GV3101-mediated transformation using the uidA (β-Glucuronidase) gene as a reporter. Influence of the time of sonication and the concentrations of protein phosphatase inhibitor
(okadaic acid) and kinase inhibitor (trifluoperazine) on Agrobacterium-mediated transformation have been evaluated. A high transformation frequency was obtained after embryos were sonicated for
45–50 s, or treated with 1.5–2.0 μM okadaic acid or treated with 100–200 μM trifluoperazine, respectively. Protein phosphatase
and kinase inhibitors enhance Agrobacterium-mediated transformation in eastern white pine. A 2–3.5-fold higher rate of hygromycin-resistant callus was obtained with
an addition of 2 μM okadaic acid or 150 μM trifluoperazine or sonicated embryos for 45 s. Stable integration of the uidA gene in the plant genome of eastern white pine was confirmed by polymerase chain reaction (PCR), Southern and northern blot
analyses. These results demonstrated that a stable and enhanced transformation system has been established in eastern white
pine and this system would provide an opportunity to transfer economically important genes into this Christmas tree species.
Communicated by W. H. Wu 相似文献
19.
20.
Shijin Wu Huaxing Zhang Xiang Yu Jianmeng Chen 《Bioprocess and biosystems engineering》2009,32(6):845-852
The gene dehalA encoding a novel dichloromethane dehalogenases (DehalA), has been cloned from Bacillus circulans WZ-12 CCTCC M 207006. The open reading frame of dehalA, spanning 864 bp, encoded a 288-amino acid protein that showed 85.76% identity to the dichloromethane dehalogenases of Hyphomicrobium sp. GJ21 with several commonly conserved sequences. These sequences could not be found in putative dichloromethane (DCM)
dehalogenases reported from other bacteria and fungi. DehalA was expressed in Escherichia coli BL21 (DE3) from a pET28b(+) expression system and purified. The subunit molecular mass of the recombinant DehalA as estimated
by sodium dodecyl sulfate–polyacrylamide gel electrophoresis was approximately 33 kDa. Subsequent enzymatic characterization
revealed that DehalA was most active in a acidic pH range at 30°, which was quite different from that observed from a facultative bacterium dichloromethane dehalogenases of Methylophilus sp. strain DM11. The Michaelis–Menten constant of DCM dehalogenase was markedly lower than that of standard DCM dehalogenases. 相似文献