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[An attempt to investigate the effect of magnesium ions and vitamin C on activity of nitrate reductase of white headed cabbage].

The aim of this study was to investigate the effect of magnesium ions and vitamin C on changes in the activity of nitrate reductase of white headed cabbage. During storage of cabbage the activity of nitrate reductase was investigated by the Jaworski method, the level of magnesium by complexometronic method and the level of vitamin C by the Pijanowski method. In addition, the effect of a supplement of magnesium and vitamin C on some changes in the activity of the enzyme was tested. For this purpose, to the incubation mixture, containing the material under study, magnesium ions in the form of magnesium chloride or vitamin C were added in respective quantities. It was found that the nitrate reductase activity as well as the level of magnesium and vitamin C are decreasing towards the inside layers of headed cabbage. Statistical evaluation of the results indicates a high by significant correlation between factors under study. A supplement of magnesium ions or vitamin C to the investigated cabbage significantly increases the activity of the enzyme.

Ascorbic Acid↗

[Spectrophotometric determination of scandium,gallium and vanadium in white cabbage leaves].

Scandium, gallium and vanadium contents in plants is on the ppm level, although plants from industrial areas can show higher concentrations of these elements. In Department of Analytical Chemistry of Silesian University there have been elaborated new, sensitive, spectrophotometric methods of determination of scandium, gallium and vanadium using Chrome Azurol S (CAS) and Sterinol (ST). The aim of this study was the application of these methods in analysis of cultivated plants from polluted regions. White cabbage from Upper Silesia was chosen. Because the spectrophotometric methods are not selective, scandium, gallium and vanadium should be preliminary separated from interfering elements. The solvent reaction was applied for the isolation from main and trace components of investigated material. Tienoiltrifluoracetone solution in xylene was used for the extraction of scandium, mesithyloxide for vanadium and n-butyl acetate--for gallium. Interfering and not separated Fe(III) was isolated using the extraction with acetylacetone solution in CHCl3 in the case of scandium and the reduction to Fe(II) by ascorbic acid in the case of gallium and vanadium. Due to influence of Fe(II) on the vanadium determination, KCN was used as a masking agent directly after the reduction. Scandium, gallium and vanadium were determined in 6 independent samples of white cabbage after dry or wet mineralization and contents of these leemnets were found from calibration graphs. Obtain results were checked by the internal standard addition method and Atomic Emission Spectrometry Method (ICP AES). The amounts of gallium and vanadium in white cabbage from Upper Silesia District determined by elaborated methods are in good correlation with a literature data, although the contents of vanadium are on the toxic level. The scandium concentration is higher than in plants from not industrial areas. The standard recovery is satisfactory. The Atomic Emission Spectrometry Method gave comparable results. The proposed the spectrophotometric methods are sensitive, precise and economical too, because they require only small amounts of reagents and simple not expensive apparatus. The methods can be recommended for many laboratories to the analytical control of white cabbage and after adaptation to the other plant material analysis.

Brassica↗

Dynamics of pesticide residues in the autumn Chinese cabbage (Brassica chinensis L.) grown in open fields.

Residues of the pesticides chlorpyrifos, dimethoate, cyhalothrin, cypermethrin, fenvalerate, deltamethrin and chlorothalonil in the autumn Chinese cabbage (Brassica chinensis L.) were studied using gas chromatography. The results indicated that the residues were dependent on the frequency and rate of pesticide application and on the weather conditions immediately following spraying. When the Chinese cabbages were treated once at normal rates, the half-lives of the seven pesticides in the vegetable were 4.7, 5.3, 2.9, 3.5, 5.4, 4.3 and 4.0 days respectively. Thus, based on the recommended preharvest intervals, the residual levels of all pesticides were within the national standards. When the Chinese cabbages were treated four times at maximal rates at 5 day intervals, the half-lives of chlorpyrifos, cypermethrin and chlorothalonil in the vegetable were 3.6, 2.9 and 5.9 days respectively, so that, based on the recommended preharvest intervals, the residual levels of chlorpyrifos and chlorothalonil exceeded the national standards for pollution-free vegetables, but the residual level of cypermethrin was within the national standard. Effects of rain on pesticide final residues in plants treated four times were more significant than in those treated once.

Agriculture↗

Copper toxicity and bioaccumulation in Chinese cabbage (Brassica pekinensis Rupr.).

Copper is among the major heavy metal contaminants in the environment with various anthropogenic and natural sources. Human health risk from heavy metal bioaccumulation in vegetables has been a subject of growing concern in recent years. To investigate Cu phytotoxic effects and bioaccumulation in the popular vegetable Chinese cabbage (Brassica pekinensis Rupr) as well as the implications for human health due to Cu in the vegetable supply, seed germination and pot culture experiments with this vegetable were carried out. Six levels (0, 0.008, 0.031, 0.125, 0.5, and 2.0 mM/L) and 3 levels (0, 0.2, and 1.0 mM/kg) of Cu treatments were performed for the seed germination and pot culture experiments, respectively. The LC(50) of Cu for seed germination of Chinese cabbage was 0.348 mM/L. In the pot culture experiments, Cu treatments significantly increased electrolyte leakage and peroxidase activity of shoot tissues, demonstrating Cu phytotoxicity to the plants. On the other hand, Cu treatments significantly stimulated, instead of reduced, chlorophyll content. Cu treatments did not show a significant effect on shoot biomass. Compared to the control, Cu treatments significantly elevated the Cu content of the shoots-9.9, 42.5, and 119.0 mg/kg (DW) of Cu were detected in the 0, 0.2, and 1.0 mM/kg treatments, respectively. These results showed that although the plants accumulated an elevated copper content and suffered damage to some extent under Cu treatment, they looked healthy. It was suggested that Chinese cabbage with an elevated Cu content and without showing visible symptoms of damage possibly could cause a risk to human health from the transfer of the metal in food.

Biomass↗

Cloning and expression of a new isotype of the peroxiredoxin gene of Chinese cabbage and its comparison to 2Cys-peroxiredoxin isolated from the same plant.

A cDNA encoding a newly identified isotype of peroxiredoxin (Prx) was isolated from a Chinese cabbage flower bud cDNA library and designated CPrxII. Database searches using the predicted CPrxII amino acid sequence revealed no substantial homology to other proteins with the exception of the yeast type II Prx with which CPrxII shares 27.8% sequence identity. Recombinant CPrxII expressed in Escherichia coli was able to protect glutamine synthetase from inactivation in a metal-catalyzed oxidation system and to reduce H2O2 with electrons provided by thioredoxin. This specific antioxidant activity of CPrxII was about 6-fold higher than that of 2Cys-Prx of the same plant. In contrast to 2Cys-Prx, which is predominantly expressed in leaf tissue of cabbage seedlings, CPrxII is highly expressed in root tissue as revealed by Northern and Western blot analyses. The CPrxII gene exists as a small multigene family in the cabbage genome.

Amino Acid Sequence↗

Characterization of salicylic acid-induced genes in Chinese cabbage.

Salicylic acid is a messenger molecule in the activation of defense responses in plants. In this study, we isolated four cDNA clones representing salicylic acid-induced genes in Chinese cabbage (Brassica rapa subsp. pekinensis) by subtractive hybridization. Of the four clones, the BC5-2 clone encodes a putative glucosyltransferase protein. The BC5-3 clone is highly similar to an Arabidopsis gene encoding a putative metal-binding farnesylated protein. The BC6-1 clone is a chitinase gene with similarities to a rapeseed class IV chitinase. Class IV chitinases have deletions in the chitin-binding and catalytic domains and the BC6-1 chitinase has an additional deletion in the catalytic domain. The BCP8-1 clone is most homologous to an Arabidopsis gene that contains a tandem array of two thiJ-like sequences. These four cabbage genes were barely expressed in healthy leaves, but were strongly induced by salicylic acid and benzothiadiazole. Expression of the three genes represented by the BC5-2, BC5-3 and BCP8-1 clones were also induced by Pseudomonas syringae pv. tomato, a nonhost pathogen that elicits a hypersensitive response in Chinese cabbage. None of these four genes, however, was strongly induced by methyl jasmonate or by ethylene.

Acetates↗

Characterization of a pollen-preferential gene, BAN102, from Chinese cabbage.

We isolated and characterized a pollen-preferential gene, BAN102, from Chinese cabbage and analyzed the activity of its promoter. There were three or four copies of the BAN102 gene in the Chinese cabbage genome that specifically expressed in pollen and pollen tube. There were 2137 bp of BAN102 genomic clone comprising 186 bp of protein coding region, and 1178 bp of 5' and 773 bp of 3' non-coding regions. TATA box were located at 1071 nt of the promoter region while the polyadenylation signal and polyadenylation site were at 1470 and 1486 nt of the 3' non-coding region. BLAST search of BAN102 sequence showed that coding region of BAN102 gene was the greatest percent similarity with arabinogalactan protein (AGP23) gene from Arabidopsis thaliana. Promoter analysis using GUS gene as a reporter showed that the pollen-specificity of BAN102 resided within the -112 to -44 bp of proximal promoter from the transient expression in tobacco and Chinese cabbage plants.

Base Sequence↗

Functional analysis of a novel male fertility CYP86MF gene in Chinese cabbage (Brassica campestris L. ssp. chinensis makino).

In our earlier work, a cytochrome P450 CYP86MF gene was isolated from floral bud of Chinese cabbage (Brassica campestris L. ssp. chinensis Makino, syn. B. rapa L.) by mRNA differential display PCR (DD-PCR) and rapid amplification of cDNA ends (RACE). To unravel the biological function of CYP86MF gene, the antisense fragment from the CYP86MF gene was transferred into Chinese cabbage pak-choi (B. campestris ssp. chinensis var. communis Tsen et Lee). Out of 22 plants transformed with the antisense gene constructed from the CYP86MF, 20 reached to flowering stage. Morphological investigations showed that the transgenic plants developed the normal floral organ. However, they remained self-infertile, even when artificial self-pollination was performed in the bud stage. Pollen germination test indicated that the pollen from the transgenic line TB-2 could not germinate normally. Further physiological, biochemical and cytological analyses showed that only significant difference was detectable in contents of the endogenous hormones, and a layer of unknown material adhered to the surface of microspore. The present studies thus provided valuable clues for understanding the biological function of the CYP86C subfamily genes. Furthermore, our studies also demonstrate a novel method for obtaining artificial male sterility line of Chinese cabbage.

Base Sequence↗

Artificial synthesis of interspecific chimeras between tuber mustard (Brassica juncea) and cabbage (Brassica oleracea) and cytological analysis.

Interspecific chimeras between tuber mustard and red cabbage were obtained by in vitro graft-culture method. Before grafting, 6-day-old seedlings of tuber mustard and red cabbage were vertically half-cut and treated with different concentrations of 6-BA and NAA for 1 min, then, they were symmetrically fit together. As a result, sectorial chimeras were initially produced from the united shoot tips. The maximum frequency of chimeral bud formation reached 6.33% when the vertical sections of tuber mustard and cabbage were treated with 2 mg/l 6-BA and 1 mg/l NAA. When sectorial chimeras were propagated on MS medium containing 1 mg/l 6-BA, periclinal and mericlinal chimeras gradually developed. Chimeral shoots were rooted on half-strength MS medium containing 0.1 mg/l NAA. The rooted chimeras were acclimatized and transferred to the field for cytological and morphological analysis. The results showed that stomata density in the chimeras was significantly higher than that of their parents, while chloroplast size, starch grain size and number were intermediate between the two parents. The chimeras were further analyzed by flow cytometry, and the results indicated that they contained both sets of parental chromosomes. Moreover, chimeral plants possessed valuable characters from the two parents.

Brassica↗

Molecular and functional characterization of a cyclophilin with antifungal activity from Chinese cabbage.

An antifungal protein that inhibits the growth of filamentous fungal pathogens was isolated from Chinese cabbage (Brassica campestris L. ssp. pekinensis) by affinity chromatography on Affi-gel blue gel and ion exchange chromatography on CM-Sepharose. The N-terminal amino acid sequence of the protein was highly homologous to that of plant cyclophilins and consequently the protein was denoted as C-CyP. To understand the antifungal activity of C-CyP, we isolated a cDNA encoding its gene from a Chinese cabbage leaf cDNA library. The Chinese cabbage genome bears more than one C-CyP gene copy and C-CyP mRNA is highly expressed in all tissues except the seeds. Recombinant C-CyP catalyzed the cis-trans inter-conversion of the Ala-Pro bond of the substrate, which indicates this protein has peptidyl-prolyl cis-trans isomerase activity. It also inhibited the growth of several fungal pathogens.

Amino Acid Sequence↗

Alteration in uptake and translocation of essential nutrients in cabbage by excess lead.

To observe the tolerance limit of lead phytotoxicity in cabbage (Brassica oleracea L.) var. Golden Aker plants were grown in refined sand with complete nutrient solution for 41 days. On 42nd day, pots with plants were separated into six lots. One lot was allowed to grow as such and was treated as control, in rest of the five lots, lead was applied at 0.1, 0.2, 0.4, 0.5 and 1.0 mM as lead nitrate. At d 75 (34 days after metal exposure), the lead toxicity symptoms as restricted growth was observed on plants at 1.0 mM lead supply. Excess lead (0.5 and 1.0 mM) developed interveinal chlorosis along the margins of young leaves. The affected leaves were reduced in size giving plant a rosette like appearance. Head size was markedly reduced at these (0.5 and 1.0 mM) levels of lead. At 0.5 mM the intensity of symptoms was markedly low. With an increase in lead supply, the concentration of lead and zinc was increased whereas that of P, S, Fe, Mn and Cu were decreased in various parts of cabbage. At 1.0 mM Pb, the concentration of lead was highest in roots and lowest in head. In leaves of cabbage the threshold of toxicity and toxicity values were 150 and 320 microg g-1 dry matter, respectively.

Brassica↗

Plant-derived biomolecules in fermented cabbage.

The formation of plant-derived biomolecules during sauerkraut fermentation was studied. Cabbage was fermented with a starter culture, and the results were compared to the results of spontaneous fermentation. The concentration of flavonoids and glucosinolates was analyzed by HPLC, and that of the glucosinolate breakdown products, by GC-MS. Of the 20 different flavonoids tested, only kaempferol was found (0.9 mg/ kg FW, fresh weight). The content of kaempferol remained constant in the cabbage fiber matrix over the fermentation process. The nitrite concentration was below the detection limit in both fermentations. The total glucosinolate content in the raw material was 3.71 micro mol/g DW, dry weight. Glucosinolates were totally decomposed in both fermentations during two weeks, and different types of breakdown products were formed. Isothiocyanates, indole-3-carbinol, goitrin, allyl cyanide, and nitriles were determined in the fermented cabbage. Isothiocyanates and allyl cyanide were the predominant breakdown products in both fermentations. Sulforaphane nitrile and goitrin were found only in small quantities in the end products.

Brassica↗

Molecular cloning, expression, and functional characterization of a 2Cys-peroxiredoxin in Chinese cabbage.

A cDNA (C2C-Prx) corresponding to a 2Cys-peroxiredoxin (2Cys-Prx) was isolated from a leaf cDNA library of Chinese cabbage. The predicted amino acid sequence of C2C-Prx has 2 conserved cysteines and several peptide domains present in most of the 2Cys-Prx subfamily members. It shows the highest sequence homology to the 2Cys-Prx enzymes of spinach (88%) and Arabidopsis (86%). Southern analysis using the cDNA insert of C2C-Prx revealed that it consists of a small multigene family in Chinese cabbage genome. RNA blot analysis showed that the gene was predominantly expressed in the leaf tissue of Chinese cabbage seedlings, but the mRNA was generally expressed in most tissues of mature plant, except roots. The expression of C2C-Prx was slightly induced by treatment with H2O2 (100 microM) or Fe3+/O2/DTT oxidation system, but not by ABA (50 microM) or GA3 (10 microM). The C2C-Prx is encoded as a preprotein of 273 amino acids containing a putative chloroplast-targeting signal of 65 amino acids at its N-terminus. The N-terminally truncated recombinant protein (deltaC2C-Prx) migrates as a dimer in a non-reducing SDS-polyacrylamide gel and as a monomer in a reducing condition. The deltaC2C-Prx shows no immuno cross-reactivity to antiserum of the yeast thiol-specific antioxidant protein, and vice versa. The deltaC2C-Prx prevents the inactivation of glutamine synthetase and the DNA cleavage in the metal-catalyzed oxidation system. In the yeast thioredoxin system containing thioredoxin reductase, thioredoxin, and NADPH, the deltaC2C-Prx exhibits peroxidase activity on H2O2.

Amino Acid Sequence↗

Effect of spraying selected pesticides on the contents of specified minerals in cabbage.

The present study was conducted to assess the effect of spraying of pesticides on the content of minerals in cabbage harvested at varying waiting periods (0, 7, 14 and 21 days) after spraying. The amounts of phosphorus and zinc in cabbage were significantly (p < 0.05) decreased while those of iron, calcium and potassium were significantly (p < 0.05) increased by the sprayings of pesticides. At varying waiting periods after the spraying of pesticides calcium, phosphorus, zinc and potassium contents of cabbage were found to be decreased remarkably (p < 0.05) while the content of iron was increased significantly (p < 0.05). Between the two pesticides, the spraying of endosulfan exerted a more pronounced effect in altering the contents of minerals than the spraying of malathion.

Brassica↗

Molecular characters and morphological genetics of CAL gene in Chinese cabbage.

BcpCAL, the homologous gene of CAL, was isolated from Chinese cabbage. Unlike BobCAL of cauliflower, BcpCAL did not hold the terminating mutation in the fifth exon. After crosses of cauliflower with Chinese cabbage, the resultant hybrids failed to form curd, which implicates the genetic complement of BcpCAL to the mutated BobCAL in the function of curd formation. One of CAL gene isolated from the hybrid apparently comes from the female parent (Chinese cabbage) even though there are a few of the bases substituted and deleted. The result offers the molecular and genetic evidences for the study of biological function of CAL in morphological genetics of curd.

Base Sequence↗

Expression of biologically active human butyrylcholinesterase in the cabbage looper (Trichoplusia ni).

This investigation examined the utility of three recombinant protein-expression systems (COS cells, insect cells and insect larvae) to cost-effectively produce biologically active human butyrylcholinesterase (BuChE). It was determined that baculovirus-infected insect cells (Sf9 and High 5) expressed 3.5- and 8.2-fold, respectively, more active enzyme than COS-7 cells. Baculovirus-infected cabbage looper (Trichoplusia ni) insect larvae produced over 26 times more than High 5 cells; in fact, one baculovirus-infected insect larva provided more active protein than 100 ml of insect cell culture. Analysis of the larvally expressed proteins revealed that the vast majority of BuChE expressed was inactive due to extensive degradation that occurred in vivo. However, the active form of BuChE does have enzyme kinetics similar to those of its human serum counterpart. Cabbage looper larvae were also examined for their ability to serve as an in vivo animal model to study protection against anti-cholinesterase toxicity. This was unsuccessful due to their high tolerance to the very toxic organophosphorus compounds tested. This tolerance may be attributed at least in part to a novel endogenous organophosphorus acid anhydride hydrolase activity that is capable of hydrolysing the chemical-warfare nerve agents sarin (isopropyl methylphosphonofluoridate) and soman (pinacolyl methylphosphonofluoridate). These results show that cabbage looper larvae can serve as an inexpensive recombinant protein-expression system for human BuChE.

Animals↗

The symbiosis of Bacillus subtilis L-forms with Chinese cabbage seedlings inhibits conidial germination of Botrytis cinerea.

AIMS: To establish whether germination of Botrytis cinerea was affected by the symbiosis of Bacillus subtilis L-form bacteria with Chinese cabbage. METHODS AND RESULTS: Germinating seeds of Chinese cabbage were co-cultivated with either L-forms of Bacillus subtilis or 5% (w/v) mannitol by soaking for 3 h. Seeds were then washed in sterile water, sown on a minimal medium and incubated in controlled conditions. L-form symbiosis was detected over a time course by ELISA. Conidial germination of Botrytis cinerea was significantly reduced on cotyledonous leaves of L-form-treated plants compared with controls. CONCLUSIONS: Symbiosis of B. subtilis L-form bacteria during seed germination of Chinese cabbage inhibits conidial germination in plants on subsequent exposure to Botrytis cinerea. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first account of plant symbiosis with L-form bacteria showing antagonism to a fungal plant pathogen. This has promising implications for the use of this L-form as a biocontrol agent.

Bacillus subtilis↗

cDNA cloning and characterization of enolase from Chinese cabbage, Brassica campestris ssp. Pekinensis.

An enolase-encoding cDNA clone from Chinese cabbage, Brassica campestris ssp. Pekinensis, was isolated. This gene (Accession number: AY307448) had a total length of 1580bp with an open reading frame of 1335bp, and encoded a predicted polypeptide of 444 amino acids with a molecular weight of 47.38 kDa. The deduced amino acid (aa) sequence shared identity with a number of enolases ranging from Bacillus subtilis to human beings and had much higher identity with other plant enolases than with enolases from Bacillus, yeast and human beings. Comparison of its primary structure with those of other enolases revealed the presence of an insertion of 5 amino acids in enolase of Chinese cabbage. Expression of the cloned enolase gene decreased under salt stress, but increased in response to low temperature. Southern blot analysis of genomic DNA indicated that low-copies of enolase gene were present in the genome of Chinese cabbage.

Amino Acid Sequence↗