To use Agrobacterium rhizogenes-induced hairy roots to create new germplasm of Dianthus caryophyllus, we transformed D. caryophyllus with A. rhizogenes by leaf disc for plant regeneration from hairy roots. The white hairy roots could be induced from the basal surface of leaf explants of D. caryophyllus 12 days after inoculation with A. rhizogenes ATCC15834. The percentage of the rooting leaf explants was about 90% 21 days after inoculation. The hairy roots could grow rapidly and autonomously in liquid or solid phytohormone-free MS medium. The transformation was confirmed by PCR amplification of rol gene of Ri plasmid and silica gel thin-layer chromatography of opines from D. caryophyllus hairy roots. Hairy roots could form light green callus after cultured on MS+6-BA 1.0-3.0 mg/L + NAA 0.1-0.2 mg/L for 15 days. The optimum medium for adventitious shoots formation was MS + 6-BA 2.0 mg/L + NAA 0.02 mg/L, where the rate of adventitious shoot induction was 100% after cultured for 6 weeks. The mean number of adventitious shoot per callus was 30-40. The adventitious shoots can form roots when cultured on phytohormone-free 1/2 MS or 1/2 MS +0.5 mg/L NAA for 10 days. When the rooted plantlets transplanted in the substrate mixed with perlite sand and peat (volume ratio of 1:2), the survival rate was above 95%.
Agrobacterium ; Chromatography, Thin Layer ; Culture Media ; Dianthus ; growth & development ; Plant Growth Regulators ; Plant Leaves ; Plant Roots ; growth & development ; Plants, Genetically Modified ; Rhizobium ; Tissue Culture Techniques ; Transformation, Genetic

Objective To establish antigen capture ELISA methed to detect Kaposi's sarcoma-as-sociated herpesvirus(KSHV)antigen,and to evaluate its feasibility for clinical application.Methods The BALB/c mice and New Zealand white rabbits were injected with purified recombinant KSHV gpK8.1 proteins to prepare the monoclonal antibody(McAb)and polyclonal antibody(PcAb)anti-gpK8.1,respectively.A new antigen capture ELISA method was established for KSHV antigen detection.The detection reproducibili-ty as well as the sensitivity and specificity of this new assay were determined by the optimization test,which antibody pairs were analyzed to choose the best coating antibody and detecting antibody.The 3 KSHV posi-tive patients sera and 257 patients sera from sexually transmitted disease,cancers or gynecological diseases were detected with this assay to evaluate its value for clinical application.Results When the McAb as coat-ing antibody at concentration of 5 μg/ml and PcAb as detecting antibody at concentration of 1.6μg/ml were selected,the highest P/N value could be obtained.The sensitive analysis of this test could detect recombi-nant KSHV gpK8.1 antigen of 31.28 ng/ml.Meanwhile,it is highly specific to detect KSHV antigen with-out cross reaction to Epstein-Barr vims(EBV),herpes simplex virus(HSV)-1 or HSV-2.All of three KSHV-positive sera and 4 sera from 257 clinical samples were positive with this new assay.which indicated that it could be used for capturing KSHV antigen.Conclusion A sensitive and specific McAb-based anti-gen capture ELISA method to detect KSHV antigen were established successfully.It is of great potential val-ue to develop reagent for KSHV clinical serologic dingnosis.

Objective To explore the clinical efficacy and safety of umbilical cord derived mesenchymal stem cells transplantation(UC-MSCT)for patients with refractory systemic lupus erythematosus (SLE).Methods Twelve patients with refractory SLE were enrolled in this study.UC-MSCs(≥106/kg cell number)were infused intravenously for each patient. The clinical manifestations and laboratory parameters were compared before and after MSCT. Results The twelve patients were followed up for one to twenty-six months after MSCT.The systemic lupus erythematosus disease activity index(SLEDAI)score decreased from 18±4 to 10±4 one month after MSCT(n=12,P＜0.01)and then decreased to 7±4 at three month follow-up.Nine patients showed improvement of 24 h proteinuria[(2103±749)mg vs(3359±1248)mg,P＜0.01]one month after MSCT.Further improvement of 24 h proteinuria was observed in eight patients[(1427±616)mg vs(3342±1333)mg,P＜0.01]at three months post MSCT.Serum creatinine of five patients decreased significantly and ten patients showed an increase of serum albumin. Serum complement C3 increased in three patients and four patients showed obvious amelioration of hematological abnormalities. There was no transplantation related complications for all the patients. Conclusion UC-MSCT is effective and safe for refractory SLE,but further observation is required to evaluate its long term efficacy.

Objective:To explore the effect of mind map on health education of children with asthma and their parents.Methods:Ninety children with asthma admitted for the first time were divided into observation group (45 cases) and control group (45 cases) by random digits table method. The control group used the traditional health education method, and the observation group used the mind map pattern health education method. The drug compliance of the two groups was evaluated before discharge and at the 1st, 2nd,3rd month after discharge, and the satisfaction of the children and parents and therapeutic effect of the two groups were compared at the time of discharge.Results:There was no significant difference in treatment adherence between the two groups before discharge ( P > 0.05). The treatment adherence at the 1st, 2nd,3rd month after discharge was (7.22±1.31), (7.58±1.55), (7.78±2.11) points in the observation group, and (6.98±2.01), (7.01±1.58), (7.11±2.48) points in the control group, there were significant differences between the two groups ( t values were 4.426, 3.884, 5.233, all P < 0.05). The satisfaction of the children and parents was (96.01±3.45) points in the observation group,and (78.13±2.39) points in the control group, there was significant difference between the two groups ( t value was 38.223, P < 0.01). The total effective rate was 77.8%(35/45) in the observation group,and 64.4%(29/45) in the control group, there was significant difference between the two groups ( χ2 value was 4.013, P < 0.05). Conclusions:Health education integrated with mind map can effectively improve the satisfaction of children and their parents, and improve the drug adherence and therapeutic effect of children.

BACKGROUND: The addition of growth factiors is commonly applied to improve the osteogenic differentiation of stem cells. However, for human pluripotent stem cells (hPSCs), their complex differentiation processes result in the unknown effect at different stages. In this study, we focused on the widely used bone forming peptide-1 (BFP-1) and investigated the effect and mechanisms of its addition on the osteogenic induction of hPSCs as a function of the supplementation period. METHODS: Monolayer-cultured hPSCs were cultured in osteogenic induction medium for 28 days, and the effect of BFP-1 peptide addition at varying weeks was examined. After differentiation for varying days (0, 7, 14, 21 and 28), the differentiation efficiency was determined by RT–PCR, flow cytometry, immunofluorescence, and alizarin red staining assays. Moreover, the expression of marker genes related to germ layers and epithelial-mesenchymal transition (EMT) was investigated at day 7. RESULTS: Peptide treatment during the first week promoted the generation of mesoderm cells and mesenchymal-like cells from hiPSCs. Then, the upregulated expression of osteogenesis marker genes/proteins was detected in both hESCs and hiPSCs during subsequent inductions with BFP-1 peptide treatment. Fortunately, further experimental design confirmed that treating the BFP-1 peptide during 7–21 days showed even better performance for hESCs but was ineffective for hiPSCs. CONCLUSION The differentiation efficiency of cells could be improved by determining the optimal treatment period.Our study has great value in maximizing the differentiation of hPSCs by adding osteogenesis peptides based on the revealed mechanisms and promoting the application of hPSCs in bone tissue regeneration.