This research uses six Agrobacterium rhizogenes R1601, R15384, R1000, A4, R1025 and R1 to infect silymarin explants to induce hairy roots and silibin. All of the six A. rhizogenes can induce Silybum marianum to generate hairy roots and the A. rhizogene A4 shows comparatively high infection on the plant. This research determines the condition to induce silymarin hairy roots by the factors of infection time, pre-culturing, co-culturing and pH value. The fact that MS liquid medium fits the proliferation of silymarin hairy roots is determined. Through PCR molecular identification, it can be seen that the DNA plasmids in the A. rhizogenes are successfully integrated into the genome of transformed roots. Using liquid chromatography, it is determined that the silibin content in silymarin hairy roots is 2.5 times that in the plant In this research, the silymarin hairy roots culturing system is established, which lays a foundation for the study of culturing silymarin hairy roots and producing silibin.
Agrobacterium ; genetics ; physiology ; Cell Culture Techniques ; methods ; Milk Thistle ; chemistry ; genetics ; growth & development ; microbiology ; Plant Roots ; chemistry ; genetics ; growth & development ; microbiology ; Silymarin ; analysis ; Transformation, Genetic

Objective:To observe the effect of rapid point pressure on clinical effect and pulmonary function of patients with chronic persistent bronchial asthma.
Methods:A total of 60 confirmed chronic persistent bronchial asthma cases were randomly allocated into two groups, 30 in each group. Cases in the treatment group were treated with rapid point pressure, 1 h for the initial treatment, and 40 min for the ensuing treatments. The treatment was done once a day for 40 d. Cases in the control group were treated with Compound Methoxyphenamine Hydrochloride Capsules, 2 capsules for each dose, 3 times a day. The treatment lasted for 7 consecutive days. A 1-year follow-up was made for both groups.
Results:After treatment, the scores of each symptom and total symptom scores for the traditional Chinese medicine (TCM) symptoms in the treatment group were significantly higher than those in the control group (P<0.05);the total effective rates in the treatment group were higher than those in the control group in each time frame; and there was a between-group statistical significance in total effective rate after 7 d of treatment (P<0.01). After 40-day treatments, there were statistical significances in the forced expiratory volume in 1 s percentage of predicted value (FEV 1%) and inspiratory capacity (IC%) in the treatment group (P<0.05); however, there were no statistical significances (P>0.05) in forced expiratory volume in 1 second to forced vital capacity ratio (FEV1/FVC), peak expiratory flow rate (PEF%) and maximum midexpiratory flow rate (MMEF%); and there were no statistical significances in pulmonary function parameters in the control group. In addition, as for pulmonary function parameters, there were no intra-group statistical significances in differences before and after treatment (P>0.05).
Conclusion:Rapid point pressure can alleviate patients’ TCM symptoms and improve their FEV%and IC%.

The effect of initial substrate concentration on the growth,metabolic activities of Photosynthetic bacteria (PSB) in the process of hydrogen production is studied.The empirical relation of the initial substrate concentration to the specific growth rate,specific substrate consumption rate and specific hydrogen production rate of PSB are developed based on the modified Monod model.It is found that the results given by the model are well agreed with the experimental data in growth phase and stationary phase of PSB.Meanwhile,the optimal substrate concentration in the process of growth,metabolism and hydrogen production is 50 mmol/L.Furthermore,when the initial substrate concentration deviates 50 mmol/L,the activation of PSB is limited or inhibited,and the inhibiting effect is more prominent than the limiting effect.In addition,it is demonstrated that initial substrate concentration has little effect on the specific substrate consumption rate.

Objective To investigate the expression of caspase-10 in differentiated thyroid carcinoma and association with its development and metastasis. Methods Thyroid samples from 37 patients in a period from January 2006 to December 2007, with differentiated thyroid carcinoma were retrospectively analyzed for caspase-10 by immunohistocbemistry(streptavidin-perosidase, S-P), compared to control group of 46 cases with nodtdar goiter. The relationship between the expression of caspase-10 and the clinical pathologic characteristics of thyroid carcinoma were also explored simultaneously. Results caspase-10 were observed as brown or yellow particles located in the cytoplasm or cell membrane of nodular goiter but there were no significant evidence for its positive expression in thyroid carcinoma, caspase-10 expression was markedly down-regulated in differentiated thyroid carcinoma(29.73%,11/37) compared with benign nodules(71.74%,33/46, χ2=14.528, P<0.01). The positive expression in 18 cases with lymph node metastasis(11.11%,2/18) was significantly lower than those in 19 patients without lymph node metastasis(47.37%,9/19; χ2=4.210, P<0.01). There was no significant correlation(P> 0.05) between the expression of caspase-10 and the clinical pathologic characteristics including male, age, TNM stage and pathologic type. Conclusion Down-regulation of caspase-10 may play a critical role in carcinogenesis and development of differentiated thyroid carcinoma.

BACKGROUND: In the past, the incidence rates of infectious diseases,natural epidemic diseases and endemic diseases in three gorges areas were always high. Because the flooded areas of Kaixian was enormous, the population of Kaixian was intensive density, the pollution may harm to the population health after the dam has been established.OBJECTIVE: To predict the influence of the Three-gorges dam on population health in Kaixian County of Chongqing and put forward to scientific evidence of preventive measures through analyzing the main health indicators and influence factors in Kaixian.DESIGN: Cross-sectional study.SETTING: Public Health College of Chongqing Medical University and Disease Controlled Center in Kaixian County of Chongqing.PARTICIPANTS: The survey was conducted in Kai County of Chongqing,from February to May 2004. All disease surveillance data of people in this county was investigated and other people lived in three-gorges area or around area were selected as the controls.METHODS: Collect and sort relevant indicators of social development,population health and public health system of Kaixian and related area to go deep into the representative townships to do some filed investigation,collect water samples of local drainage areas to carry out laboratory quantitative analysis; carry out comparative analysis based on the relevant materials; carry out comprehensive predicting and evaluation to the potential impact on population health of the three gorges engineering by combining the quantitative and qualitative analysis methods.general people in Kai county, Chongqing city and all over the nations durKai county, Chongqing city and all over the nations during 1999 and 2003;ty, Chongqing city and all over the nations during 1999 and 2001: Mortality rates of infants and under-five children were also higher than that average levels of whole country and Chongqing respectively; Multi-indicators showed that the general population health status of Kaixian was great bad,which stood a distinct gap among average levels of the whole country and Chongqing maintained about 250/100 000 or so, which was higher than the national level (190/10 000), however, the incidence rate of infectious diseases of Kaixian was rising year after year, even that of the recent two years has been reached up to more than 1.5 times higher than that of the whole of the water-based infectious diseases, especially the virus hepatitis, was severe. Another potential impact on population health of three-gorge dam is the emergence of natural epidemic diseases and insect-based infectious diseases. Malignant tumor such as hepatocarcinoma.CONCLUSION: We should strengthen current public health system, and cognition to the possible threat on disease epidemic and public health.With the construction of three-gorge dam, it is necessary to speed up establishment of effective measures to reduce predictable influence on population health.

To improve ethanol production in Saccharomyces cerevisiae,an integration plasmid pUPGKAT with PGK promoter(phosphoglycerate kinase promoter),adh1 gene(the coding sequences of alcohol dehydrogenaseⅠ) and CYC1 terminator(Cytochrome c transcription terminator) was constructed.Firstly,a fusion fragment composed of PGK promoter and adh1 gene was generated by over lap extension PCR and ligated into pUG6 resulting in plasmid pUPGKA.Subsequently,CYC1 termi nator was amplified from pSH65 by PCR and ligated to the SpeⅠand SacⅡrestriction site of pUPGKA.To integrate PGK-adh1-CYC1 into S.cerevisiae genome,pUPGKAT was digested by TthⅢⅠand the lin-earized plasmid was used to transform S.cerevisiae YS2-△adh2(adh2 disrupted strain) by lithium acetate method.The yeast mutant YS2-△adh2-adh1 which had the adh1 gene placed under the PGK promoter and harbored the adh2 deletion was constructed.Anaerobic fermentation showed overexpression of adh1 by PGK promoter resulted in a 8.84% higher ethanol production compared to YS2-△adh2.

Acyl carrier protein is an essential component involved in the biosynthesis of DHA(Docosahexaenoic Acid) via PKS(Polyketide synthase) pathway,which takes the growing acyl chain from one enzyme to another.One cDNA clone,with high homology of ACP,was isolated from Schizochytrium sp.FJU-512 cDNA library.The deduced amino acid sequence contained 142 residues with isoelectric point of 5.04 and had the 4'-phosphopantetheine prosthetic(4'-PP) binding site.The target fragment was digested with BamHⅠ/HindⅢand inserted into the expression vector pET-30a resulting in the plasmid pET-30a/acp.The recombinant vector was transformed into E.coli BL21(DE3) and induced by IPTG.SDS-PAGE analysis demonstrated that ACP was effectively expressed.

Objective To construct the recombinant plasmid pYES6-S and express and purify spike protein of severe acute respiratory syndrome(SANS)coronavirus in Saccharomyces cerevisiae. Methods DNA fragments of SANS coronavirus were obtained by reverse transeription.Four over- lapped fragments of spike protein genes were amplified by polymerase chain reaction(PCR)and ligated into an integral spike protein gene by restriction enzyme digestion.The spike protein gene recombined with pYES6 and cloned into E.coll.The recombinant plasmid pYES6-S was induced and expressed in Saccharomyces cerevisiae(INVScl)by galactose.Results The recombinant plasmid pYES6-S was confirmed that inserted fragment was right in length,direction and base matching by restriction enzyme digestion and sequencing.The purified protein encoded by the whole spike protein gene was about Mr 110?10~3 identified by electrophoresis.Conclusion The whole spike protein gene of SARS coronavirus is cloned into E.coli and the protein is expressed in Saccharomyces cerevisiae successful ly.which can be helpful in SARS vaccine research.

The rate of corneal graft rejection is still high for high-risk keratoplasty although immune suppression drug is routinely used.The role of traditional Chinese medicine in corneal transplantation is concerned gradually.Heijingpaichitang on the prevention and treatment of rats with high-risk corneal allograft rejection needs further study.Objective This study was to investigate the inhibitory effect of heijingpaichitang on high-risk corneal transplantation immune rejection in rats.Methods Sixteen female SD rats were used as the donors and 32female Wistar rats were served as recipients.The high-risk corneal trasplantation models were established by corneal suture in 32 Wistar rats,and then homogeneity variant SD-Wistar corneal transplantation was performed.The recipients were randomized into model control group,cyclosporinc A (CsA)group,heijingpaichitang group and CsA +heijingpaichitang group.CsA,heijingpaichitang and CsA + heijingpaichitang was orally administered 4 days after operation once per day for 15 days,and normal saline solution was used at the same way in the model control group.Ocular anterior segment reaction was examined under the slit lamp and corneal opacification,edema and neovasculation were scored based on Larkin' s criteria.Rejection index of the corneal graft was recorded and the graft survival time was calculated.The pathological examination of the corneal graft was carried out in all rats,and the inflammatory cells in the corneas and CD4+ cells in the periphery blood were assayed using flow cytometry.The use of the animals complied with ARVO Statement.Results Corneal graft rejection occurred in 10 days after operation in the model control group,12-13 days in the CsA group and heijingpaichitang group and 22 days in the CsA +heijingpaichitang group.Compared with model control group,the scores of the corneal opacification,corneal edema and neovascularization were significantly lower in the CsA group,heijingpaichitang group and CsA+heijingpaichitang group (P＜0.05),and all the scores were declined in the CsA+ heijingpaichitang group compared with CsA group and heijingpaichitang group(P＜0.01),but no significant differences were seen in the scores between the CsA group and heijingpaichitang group(P＞0.05).The mean survival time of grafts was (10.38 ±1.69)days in the model control group,(22.50 ± 3.07) days in the CsA + heijingpaichitang group,with the significant difference (t =-9.790,P =0.000).The pathological examination of graft showed that the lymphocytes and new blood vessels were less in the CsA+heijingpaichitang group compared with CsA group and heijingpaichitang group 15 days after operation.Flow cytometry verified that the number of lymphocytes in graft,CD4+cells and CD4+/CD8+ in periphery blood were significantly lower in the heijingpaichitang group,CsA group and CsA+heijingpaichitang group compared with model control group (P＜0.05).Conclusions Heijingpaichitang can inhibit immune rejection to certain extent in high-risk corneal transplantation rat and has a similar effect to 0.1％ CsA.Heijingpaichitang and 0.1％ CsA have a synergistic effect.

Objective To explore the clinical characteristics and the cause of misdiagnosis of child suffering from cerebral parago-nimiasis with intracranial hemorrhage as initial symptom,and to improve the diagnosis and treatment level of cerebral paragonimiasis.Methods The clinical data of the children who suffered from cerebral paragonimiasis with intracranial hemorrhage as initial symptom were collected from January 2011 to December 2015 in Affiliated Hospital of Zunyi Medical College.The clinical manifestation,imageology and laboratory tests,outcome of therapy were analyzed and then the effect of treatment and the prognosis were followed up.Results There were 7 patients meeting the inclusion criteria for cerebral paragonimiasis,including 4 male and 3 female.They were from 6 to 13 years old with the average age of 9.3 years old.All patients presented with headache and vomiting,and showed intracranial hemorrhage through CT or MRI of brain.All of the 7 patients were misdiagnosed as cerebrovascular malformation by the neurosurgeons.Three of them showed typical imaging pattern including tunnel sign and the ring-like shape of cerebral paragonimiasis.Five of these cases were attacked by pulmonary distomiasis at the same time.Six of them had an increasing eosinophil,and the paragonimus antibody was positive.They were treated with Praziquantel.Six patients recovered completely,and 1 patient had the dysfunction of left extremities.Conclusions The childhood cerebral paragonimiasis has strong clinical heterogeneity and diversity in manifestation.Intracranial hemorrhage may be the initial symptom,which should be paid more attention to.Cerebral paragonimiasis can be diagnosed and treated early according to the clinical characteristics,the increase of eosinophil,the typical changes in imageology and the specific antibody test.