Production of ascorbic acid, total protein, callus and root in vitro of non-heading Chinese cabbage by tissue culture.

The objective of the present work was the selection of cultivar, suitable medium and explant type for callus, root production, ascorbic acid, total ascorbic acid, dehydroascorbic and total protein of non-heading Chinese cabbage in two cultivars 'Caixin' and 'Suzhouqing'. We compared 10 types of MS media supplemented with 0.0, 1.0, 2.0 and 3.0 mg/l TDZ; 0.0, 0.25, 0.50 and 1.0 mg/l NAA and 0.0, 5.0, 7.5 and 9.0 mg/l AgNO3 and 5 kinds of explants as embryo, leaf, root, cotyledon and hypocotyl. Maximum frequency of callus fresh weight was recorded with hypocotyl explant, which were cultured on MS + 2.0 mg/l TDZ + 1.0 mg/l NAA + 9.0 mg/l AgNO3 in 'Suzhouqing', optimum callus dry weight was obtained on the same media. The highest result for root fresh and dry weight recorded with 'Caixin' with MS + 3.0 mg/l TDZ + 1.0 mg/l NAA + 9.0 mg/l AgNO3 when we used embryo as explant. The highest ascorbic acid content was found with callus cultured on MS + 1.0 mg/l TDZ + 0.25 mg/l NAA + 5.0 mg/l AgNO3, when used leaf explant in 'Caixin' or root in 'Suzhouqing', and there were no significant difference between them. While the highest value of total AsA content was registered with callus cultured on MS + 2.0 mg/l TDZ + 0.25 mg/l NAA + 5.0 mg/l AgNO3 extracted from cotyledon in 'Caixin'. The highest content of DHA was registered with MS + 2.0 mg/l TDZ + 0.25 mg/l NAA + 5.0 mg/l AgNO3 with cotyledon in 'Caixin'. Also, in 'Caixin' MS + 3.0 mg/l TDZ + 0.25 mg/l NAA + 5.0 mg/l AgNO3 recorded the highest value of total protein content with embryo explant.

BcAP3, a MADS box gene, controls stamen development and male sterility in Pak-choi (Brassica rapa ssp. chinensis).

Stamen development is an important developmental process controlled by multiple internal and external factors. Developmental abnormalities of stamens can disrupt the structure and function of anthers, and then result in male sterility. As well known, APETELA 3 (AP3) has a clear function in regulating stamen development, which may impact in male sterility. However, the mechanisms of stamen development and male sterility controlled by AP3 are still not very clear, particular in Pak-choi (Brassica rapa ssp. chinensis). In this work, BcAP3 encoded a protein containing a MADS-box domain, which was a homolog of AtAP3, was identified in Pak-choi. Sequence alignments and phylogenetic analysis indicated that BcAP3 was highly similar to AtAP3. BcAP3 was shown to be localized to the nucleus and exhibited the potential of transcription factor. The transcript of BcAP3 was only expressed in flowers of Pak-choi, indicating that it may act in flower development. Overexpression of BcAP3 in Arabidopsis resulted in developmental abnormalities of anther wall and low vigor pollen, which were associated with the phenotype of male sterility. Expression levels of NST1 and NST2, involved in secondary wall thickening in anther walls, were significantly higher in the BcAP3-transgenic plants than in control plants, suggesting that BcAP3 may affect anther wall development by regulating NST1 and NST2. Taken together, our study demonstrated that BcAP3 could play an essential role in stamen development and male sterility.

In vitro and in vivo aphrodisiac properties of the seed extract from Allium tuberosum on corpus cavernosum smooth muscle relaxation and sexual behavior parameters in male Wistar rats.

BACKGROUND: Allium tuberosum is a well-known spice as well as a herb in traditional Chinese medicine, used for increasing libido and treating erectile dysfunction. However, not many studies have been done to evaluate the sexual enhancing properties of A. tuberosum. The aim of this study was to evaluate the aphrodisiac and vasorelaxant properties of A. tuberosum on corpus cavernosum smooth muscle (CCSM) as well as checking the effect on enhancing male rat sexual behavior, libido, potency as well as its spermatogenic properties. METHOD: The seeds were powdered and sequentially extracted with hexane, ethyl acetate and butanol. Male Wistar rats were administered with graded doses of the n-BuOH extracts (ATB) of A. tuberosum (50, 100, 200 and 400 mg/kg) and Viagra was used as the positive control drug. The extract/drug was administered by gastric probe once daily for 45 days and the sexual behavior was analyzed by exposing the male rats to female rats in the estrus period. RESULTS: ATB relaxed corpus cavernosum smooth muscle (68.9%) at a concentration of 200 µg/ml. The results obtained from the animal studies indicated that ATB significantly increased mount frequency (MF), intromission frequency (IF), ejaculation frequency (EF), ejaculation latency (EL) and markedly reduced post ejaculatory interval (PEI), mount latency (ML), and intromission latency (IL). Furthermore, a remarkable increase in the test for potency was observed as witnessed by marked increase in erections, quick flips, long flips and total reflex. In addition, ATB significantly improved the sperm viability and count as well as increased the concentrations of testosterone, follicle stimulating hormone (FSH), and phosphatases in the treated animals. CONCLUSION: Thus our results suggest that A. tuberosum could stimulate sexual arousal and enhance sexual execution in male rats, thus providing valuable experimental evidence that A. tuberosum possesses sexual enhancing properties.

Allium tuberosum: Antidiabetic and hepatoprotective activities.

Allium tuberosum (AT) is traditionally used for treating nocturnal emissions, abdominal pain, diarrhea, sexual dysfunction and asthma. This study aimed at investigating the antidiabetic and hepatoprotective activities of the butyl alcohol fraction from the methanolic extract of A. tuberosum. For the antidiabetic activity, rats were induced with diabetes by intraperitoneal injection of 150mg/kg alloxan and treated for 30days with AT extract (100, 200 and 400mg/kg). Animals were sacrificed after the study and the fasting blood glucose (FBG), triglyceride (TG), total cholesterol (TC), HDL, malondialdehyde (MDA) catalase, superoxide dismutase and glutathione levels were determined. The hepatoprotective assay, mice were pretreated for seven days with AT (100, 200 and 400mg/kg) and silymarin (100mg/kg or). Thereafter 10ml/kg of 2% v/v CCl4 was administered intraperitoneally on the 7th day to induce acute liver injury. Blood and liver samples were obtained and serum enzymes ALT, AST, ALP, SOD, GSH, CAT, MDA and pro-inflammatory mediators were assessed. AT significantly decrease FBG, serum TG, TC, MDA levels and significant increased HDL, SOD, GSH and CAT activities in the diabetic rats. In addition, AT significantly inhibited MDA, IL-1b, IL-6 and TNF-α levels and prevented the depletion of the antioxidant enzymes GSH, SOD and CAT activities in CCl4 induced liver damage. Furthermore, AT markedly reduced AST, ALT and ALP levels in the CCl4 treated mice groups. In conclusion, the antidiabetic and hepatoprotective effect of AT may be associated with its antioxidant and its ability to inhibit the pro-inflammatory mediators.

Basic helix-loop-helix transcription factor BcbHLHpol functions as a positive regulator of pollen development in non-heading Chinese cabbage.

Cytoplasmic male sterility (CMS) is a common trait in higher plants, and several transcription factors regulate pollen development. Previously, we obtained a basic helix-loop-helix transcription factor, BcbHLHpol, via suppression subtractive hybridization in non-heading Chinese cabbage. However, the regulatory function of BcbHLHpol during anther and pollen development remains unclear. In this study, BcbHLHpol was cloned, and its tissue-specific expression profile was analyzed. The results of real-time polymerase chain reaction showed that BcbHLHpol was highly expressed in maintainer buds and that the transcripts of BcbHLHpol significantly decreased in the buds of pol CMS. A virus-induced gene silencing vector that targets BcbHLHpol was constructed and transformed into Brassica campestris plants to further explore the function of BcbHLHpol. Male sterility and short stature were observed in BcbHLHpol-silenced plants. The degradation of tapetal cells was inhibited in BcbHLHpol-silenced plants, and nutrients were insufficiently supplied to the microspore. These phenomena resulted in pollen abortion. This result indicates that BcbHLHpol functions as a positive regulator in pollen development. Yeast two-hybrid and bimolecular fluorescence complementation assays revealed that BcbHLHpol interacted with BcSKP1 in the nucleus. This finding suggests that BcbHLHpol and BcSKP1 are positive coordinating regulators of pollen development. Quantitative real-time PCR indicated that BcbHLHpol and BcSKP1 can be induced at low temperatures. Thus, we propose that BcbHLHpol is necessary for meiosis. This study provides insights into the regulatory functions of the BcbHLHpol network during anther development.

Microtubule and male sterility in a gene-cytoplasmic male sterile line of non-heading Chinese cabbage.

BACKGROUND: Microtubules are the basic components of the cytoskeleton in eukaryotic cells and are made up of 13 parallel protofilaments, each composed of α- and ß-tubulin unit molecules aligned along the longitudinal axis of the microtubule. RESULTS: α-Tubulin gene TUBA2 from non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino) was expressed at the highest level in stamens and at lower levels in other organs. In addition, it was expressed at a much lower level in the cytoplasmic male sterile (CMS) line than in the maintainer line. Furthermore, at the microsporocyte stage of development in the CMS line the microtubule bundles were knitted together in random organisation, which differed significantly from the radiate microtubule bundles running circumferentially around the nucleus in the maintainer line. Also, large vacuoles appeared within the cytoplasm in the CMS line with no dyed microtubules. CONCLUSION: TUBA2 was very important to pollen development, which might be closely related to male sterility. Large vacuoles might replace the nuclei close to the cell walls and lead to a lack of microtubules when the cells abort. Abnormalities and defects in the organisation and composition of microtubules in the male sterile line highlighted the complex interaction between microtubules and cytoplasmic male sterility.

Molecular cloning and characterisation of cytoplasmic glutamine synthetase gene BcGS1 from non-heading Chinese cabbage.

BACKGROUND: Glutamine synthetase (GS; EC 6.3.1.2) is a key enzyme of nitrogen (N) assimilation, catalysing the synthesis of glutamine from ammonium and glutamate. Plants have two types of GS isoenzyme that are localised in different compartments: one in the cytosol (GS1) and the other in the chloroplast (GS2). GS1 is the major form of GS in plant roots and directly converts ammonium taken up by plant roots to glutamine. RESULTS: The GS1 gene cDNA of non-heading Chinese cabbage (Brassica campestrisssp. chinensis Makino) cultivar 'Suzhouqing' was isolated by RT-PCR (real-time polymerase chain reaction) and (5'/3')-RACE (rapid amplification of cDNA ends) techniques. It was classified as GS1 by sequence alignment and motif search and named B. campestris ssp. chinensis Makino GS1 (BcGS1). Subcellular localisation analysis showed that BcGS1 was distributed in the cytoplasm of cells. BcGS1 was expressed in all parts, but mainly in the roots, which was verified by northern blotting analysis. Additionally, its expression was influenced by the N source concentration. CONCLUSION: These results suggest that BcGS1 is a novel member of the GS family in plants. BcGS1 was significantly related to N assimilation in non-heading Chinese cabbage, demonstrating that this gene plays an important role in plant growth and development.