Research on novel pullulanase has major significance on the domestic industrialization of pullulanase and the breakdown of foreign monopoly. A thermophilic bacteria LM 18-11 producing thermostable pullulanase was isolated from Lunma hot springs of Yunnan province. It was identified as Anoxybacillus sp. by 16S rDNA phylogenetic analysis. Full-length pullulanase gene was cloned from Anoxybacillus sp. LM18-11. The optimum temperature of the pullulanase was between 55 and 60 degrees C with a half-life as long as 48 h at 60 degrees C; and its optimum pH was between 5.6 and 6.4. V(max) and K(m) of the pullulanase was measured as 750 U/mg and 1.47 mg/mL, which is the highest specific activity reported so far. The pullulanase crystals structure showed a typical alpha-amylase family structure. The N-terminal has a special substrate binding domain. Activity and substrate binding were decreased when the domain was deleted, the V(max) and K(m) were 324 U/mg and 1.95 mg/mL, respectively. The pullulanase was highly heterologous expressed in Bacillus subtilis by P43 promoter. The extracellular enzyme activity was 42 U/mL, which increased more than 40 times compared to the initial strain. This pullulanase has good application prospects.
Anoxybacillus ; classification ; enzymology ; China ; Glycoside Hydrolases ; metabolism ; Hydrogen-Ion Concentration ; Phylogeny ; RNA, Ribosomal, 16S ; genetics ; Temperature

Peking University Health Science Center has various characteristics of educational manage-ment practice such as clear three level management, diversified management training and strict quality control, and at the same time there exist many deficiencies, such as lack of the implementation of management regulations and the requirements caused by multi-level management and management quality having ups and downs be-cause of human factors and etc.. By introducing life cycle management into national continuing medical educa-tion project management, we make management run through the whole life cycle of national continuing medical education project, namely five aspects of management from reporting and publishing, planning and publicity, holding and supervision, graduation and certificates, as well as summary and feedback, which plays an impor-tant role in standardizing the national continuing medical education project and improving the quality of the project.

Objective Comparing the effects of steam sterilization on cyclic fatigue of stainless steel files. Meth-ods Fifty instruments of 25# stainless steel K files were randomly divided into 5 groups, which include 10 in each group. Group 1 was the blank control group, group 2 to 5 were subjected to 1, 3, 4, 5 steam sterilization cycles, then the files were tested by a custom-made device to assess cyclic fatigue and the number of cycles of failure (NCF) was calculated. Microstruc-ture of each file’s fracture surface was analyzed by Scanning Electron microscope (SEM). Results NCF in the 5 groups were 4 345.2 ± 384.2,3 937.9 ± 645.4,3 812.9 ± 532.5,3 626.2 ± 380.0,3 625.9 ± 565.6 respectively, and the differences among 5 groups were significantly different(F=2.598, P<0.05). After 4 or 5 times of steam sterilization, cyclic fatigue de-creased if compared with that in control group (P<0.05). Fracture surface in group 1 and group 2 was tough dimple structure and large numbers of regular, deep dimples were distributed on the surface. You could also see micro cavities clearly formed by fracture;Fracture surface in group 4 was dimple structure in brittle intergranular morphology. It is characterized by the presence of thin brittle precipitates, clean and smooth crystal interface, and a great sense of polyhedral stereo. Conclusion After 4 times of steam sterilization, cyclic fatigue strength of 25# stainless steel K files declined, which possessed the poten-tial risk of fracture.

This study aims to analyse the value of CODEHOP RT-PCR in the detection of Flavivirus. According to the amino acid sequences of polyproteins of different flaviviruses published in GenBank, a pair of primers was designed using the CODEHOP method. One-step RT-PCR was used to detect Japanese encephalitis virus strain JEV1201, Dengue virus strain JKD001, and yellow fever virus vaccine YV6161. BLAST analysis and phylogenetic analysis were performed after the RT-PCR products of nucleocapsid genes were sequenced. The results showed that this method could amplify Flavivirus specifically, and the size and sequence of the target fragment accorded with the anticipated result. JEV1201 had the highest homology to Japanese encephalitis virus strain YL2009-4/YC2009-3, belonging to the branch of the phylogenetic tree of Japanese encephalitis virus strains. JKD001 had the highest homology to Dengue virus strain DENV-2/ID/1022DN/1975, belonging to the branch of the phylogenetic tree of Dengue virus strains. YV6161 had the highest homology to Yellow fever virus strain 17D, belonging to the branch of the phylogenetic tree of Yellow fever virus strains. In conclusion, the method of CODEHOP RT-PCR can be effectively used to detect, identify, and phylogenetically analyse Flavivirus.
DNA Primers ; genetics ; Flavivirus ; classification ; genetics ; isolation & purification ; Flavivirus Infections ; virology ; Humans ; Molecular Sequence Data ; Phylogeny ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Viral Nonstructural Proteins ; genetics

Objective To investigate the roles of transforming growth factor(TGF)-β1 gene polymorphisms in severe acute respiratory syndrome-coronavirus(SARS-CoV)infection and the interstitial lung fibrosis after recovered.Methods Sixty-five recovered SARS patients,37 health care workers and 66 healthy controls were enrolled in this case-case study.The association between genetic polymorphisms of TG F-β1 and suscept ibility to SARS or interstitial lung changes after SARS reco,vered was carried out.Polymerase chain reaction-sequencing based typing(PCR-SBT)method was used to determine the polymorphisms of TGF-β1 gene at locus+869 and+915.Data were analyzed using t test and chi square test.Results There was no significant association of TGF+β1 gene polymorphisms at locus+869 and+915 in recovered SARS patients,health care workers and heahhy controls.And gene linkage of this two loci was not related with SARS-CoV susceptibility.Furthermore,no association between interstitial lung changes in recovered SARS palients and TGF-β1 gene polymorphisms or genetic linkage of this two loci.Conclusions It may not be related between TGFβ1 gene polymorphisms at locus+869 and+915 and SARS-CoV susceptibility.And interstitial lung changes in recovered SARS patients may not be influenced by TGF-β1 gene polymorphisms.

OBJECTIVETo explore the difference in the efficacy and effect mechanism of subcortical ischemic vascular disease (SIVD) complicated with depression between acupuncture and medication.

METHODSSixty patients were randomized-into an acupuncture group and a medication group, 30 cases in each one. In the acupuncture group, acupuncture was applied to Baihui (GV 20), Shuigou (GV 26), Fengfu (GV 16), Fengchi (GB 20) and the others, once a day, 6 times a week. The treatment of 4 weeks made one session and totally 2 sessions were required. In the medication group, nimodipine 30 mg, three times a day and fluoxetine 20 mg, once a day were prescribed for oral administration, for 8 weeks totally. Before treatment, at the end of the 4th week and at the end of the 8th week of treatment, cerebral blood flow velocity (CBFV) and solubility CD40 ligand (sCD40L) were determined respectively. The scores in Montreal cognitive assessment (MoCA) and Hamilton' s depression scale (HAMD) were evaluated in the two groups. The efficacies on cognitive function and depression symptoms were compared in the patients between the two groups. Results Compared with the outcome before treatment, mean blood flow velocity (Vm) of middle cerebral artery (MCA), anterior cerebral artery (ACA) and posterior cerebral artery (PCA) was increased significantly at the end of the 4th week of treatment in the two groups (all P<0.05). At the end of the 8th week, Vm was increased much significantly (all P<0.01). The differences were not significant in comparison between the two groups (all P>0.05). Compared with the expression before treatment, sCD40L was reduced significantly after treatment in the patients of the two groups (all P<0.01), but the differ- ence was not significant between the two groups (all P>0.05). Compared with that before treatment, MoCA score was increased significantly after treatment in the two groups (all P<0.01), HAMD score was reduced sig- nificantly (all P<0.01), the differences were not significant in comparison between the two groups (all P>0.05). The total effective rate of cognitive improvement was 86.7% (26/30) in the acupuncture group and was 80.0% (24/30) in the medication group, the differences were not significant in comparison of the two groups (P>0.05). The total effective rate of the improvement in depression was 93.3% (21/30) in the acupuncture group and was 86.7% (26/30) in the medication group, the differences were not significant in comparison of the two groups (P>0.05).

CONCLUSIONAcupuncture could significantly increases CBFV and reduces serum sCD40L expressions in the patients of SIVD complicated with depression, and significantly improves cognitive function and relieves depression symptoms. The efficacy of it is similar to that of western medication. The increase of serum sCD40L expression is possibly involved in the occurrence and development of SIVD. Reducing sCD40L expression contributes to the alleviation of damage induced by cerebral ischemia and reperfusion.

Acupuncture Points ; Acupuncture Therapy ; Adult ; Aged ; Cerebral Infarction ; complications ; drug therapy ; therapy ; Depression ; drug therapy ; etiology ; therapy ; Female ; Fluoxetine ; administration & dosage ; Humans ; Male ; Middle Aged ; Nimodipine ; administration & dosage ; Treatment Outcome

To explore the new method of discriminating Cistanche deserticola, Cynomorium songaricum and Orobanche pycnostachya by using PCR amplification of specific alleles. 30 samples of the different C. deserticola, 21 samples of C. songaricum and O. pycnostachya were collected. The total DNA of the samples were extracted, the ITS sequences from C. deserticola, C. songaricum and O. pycnostachya were amplified by PCR and sequenced unidirectionally. These sequences were aligned by using ClustulW. Specific primer was designed according to the ITS sequences of specific alleles, and PCR reaction system was optimized. Additionally, compare with the identification of specific PCR method and DNA sequence analysis method. The result showed that the 331 bp identification band for C. deserticola and the adulterants not amplified bands by a single PCR reaction, which showed good identification ability to the three species. PCR amplification of specific alleles can be used to identify C. deserticola, C. songaricum and O. pycnostachya successfully.
Alleles ; Cistanche ; classification ; genetics ; DNA Primers ; genetics ; DNA, Intergenic ; genetics ; DNA, Plant ; genetics ; Drug Contamination ; prevention & control ; Phylogeny ; Polymerase Chain Reaction ; methods

Experimental teaching plays an important role in pathological teaching.We have taken effective measures to renovate the methods for experimental teaching,establishing open teaching mode.As a result,teaching quality increases and students become more active in pathological learning.

OBJECTIVETo compare the clinical efficacy in the treatment of post-stroke shoulder-hand syndrome between floating-needle therapy and conventional acupuncture on the basis of rehabilitation training.

METHODSOne hundred cases of post-stroke shoulder-hand syndrome were randomized into a floating-needle group and an acupuncture group, 50 cases in each one. The passive and positive rehabilitation training was adopted in the two groups. Additionally, in the floating-needle group, the floating-needle therapy was used. The needle was inserted at the site 5 to 10 cm away from myofasical trigger point (MTrP), manipulated and scattered subcutaneously, for 2 min continuously. In the acupuncture group, the conventional acupuncture was applied at Jianqian (EX-UE), Jianyu (LI 15), Jianliao (TE 14), etc. The treatment was given once every two days, 3 times a week, and 14 days of treatment were required. The shoulder hand syndrome scale (SHSS), the short form McGill pain scale (SF-MPQ) and the modified Fugl-Meyer motor function scale (FMA) were used to evaluate the damage severity, pain and motor function of the upper limbs before and after treatment in the two groups. The clinical efficacy was compared between the two groups.

RESULTSSHSS score, SF-MPQ score and FMA score were improved significantly after treatment in the two groups (all P < 0.01), and the improvements in the floating-needle group were superior to those in the acupuncture group (all P < 0.05). The total effective rate was 94.0% (47/50) in the floating-needle group, which was better than 90.0% (45/50) in the acupuncture group (P < 0.05).

CONCLUSIONThe floating-needle therapy combined with rehabilitation training achieves a satisfactory efficacy on post-stroke shoulder-hand syndrome, which is better than the combined therapy of conventional acupuncture and rehabilitation training.

Acupuncture Therapy ; instrumentation ; methods ; Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Male ; Middle Aged ; Reflex Sympathetic Dystrophy ; etiology ; rehabilitation ; therapy ; Stroke ; complications ; Treatment Outcome ; Young Adult

Objective An analysis was conducted to investigate the iodine nutrition level of children aged 8 - 10 in low-coverage area of iodized salt of Yushu Qinghai province for providing a scientific basis for the development of effective preventive measures. Methods Yushu, Chengduo, Nangqian and Zaduo counties with higher non-iodized salt coverage rate in Yushu Qinghai province were chosen as survey counties in 2009. Three townships were selected in each county, and 2 primary schools were selected in each township and 40 urine samples of children aged 8-10 were collected randomly in each primary school. The content of urinary iodine was analyzed by As-Ce catalytic spectrophotometery. Results Median urinary iodine of children aged 8 - 10 in Nangqian and Zaduo was < 100 μg/L. The percentage of median urinary iodine < 50 μg/L in Yushu was over 20%. Median urinary iodine of children aged 10 in Zaduo was 81.5 μg/L, the percentage of median urinary iodine < 50 μg/L of children aged 9 and 10 was over 20%. The percentage of median urinary iodine < 50 μg/L in children aged 9 and 10 of Yushu was over 20%. Median urinary iodine of girls in Zaduo was 87.1 μg/L, the percentage of median urinary iodine < 50 μg/L of boys in Zaduo was over 20%. The percentage of median urinary iodine < 50 μg/L of girls in Yushu was over 20%. Conclusions The iodine nutrition level of children aged 8 - 10 in Nangqian, Zaduo and Yushu counties were very low due to the impact of non-iodized salt. We propose salt market in the region to strengthen management and improve the coverage and consumption rates of iodized salt to improve the level of iodine nutrition for effective prevention of iodine deficiency disorders.