Objective To construct a packaged bacteria strain and, with it, to prepare gene III-defective helper phage. Methods Whole gene III and chloramphenicol-resistant gene were amplified and joined to form a fragment flanking 36bp homologous sequence of hisBCD using overlapping extending PCR, and it was then integrated into chromosome of E. coli XL1-Blue to replace the hisBCD gene using Red recombination system. The recombinant strain was identified with chloramphenicol-resistant screening, PCR and histidine-phenotype analysis, and named as XL-pⅢ. The gene III-deleted genome of phage was constructed with PCR and transfected into the recombinant strain XL-pⅢ to prepare the defective helper phage. It was quantified by infecting E. coli XL1-Blue and formed colonies were counted in kanamycin plate. Results The recombinant strain could grow in chloramphenicol-resistant plate, but couldn't grow in M9 medium without histidine, implying that it had lost its histidine phenotype. The aimed gene fragment could be amplified as designed. The constructed recombinant was named as XL-pⅢ. The prepared defective helper phage could infect E. coli XL1-Blue only once and the CFU was 3.7?108. Conclusions The packaged strain XL-pⅢ is successfully constructed and used to prepare the gene III-deleted helper phage. It is expected to be used in SIP system.

BACKGROUND:Studies regarding neural stem cells (NSCs) mainly focus on rodents and human,few of which addressing non-human primate animals.OBJECTIVE:To explore a culture system of cynomolgus monkey NSCs (cmNSCs),additionally,to observe its differential potential.DESIGN,TIME AND SETTING:An in vitro cytology observation was performed at the Canter for Stem Cell Biology and Tissue Engineering,Sun Yat-sen University between July 2008 and April 2009.MATERIALS:Spontaneously aborted cynomolgus monkey fetus with 3-5 months old (gestational age),was provided by Landao Biotechnology Co.,Ltd.METHODS:The subventricular zone and the hippocampus were dissected aseptically from the brain of spontaneously aborted cynomolgus monkey fetus,then triturated with fire-polished glass Pasteur pipettes to single cells,and cultured in 25 cm~2 culture flask with concentration of (2.0-5.0)×10~5.The neurospheres were transferred into polyornithine and fibronectin precoated cell culture dish after 2 weeks when they formed.Each 20 μg/L of epithelium growth factor (EGF) and basic fibroblast growth factor (bFGF) was daily added.The adhesive cells were passagad when they reached about 90% confluency.MAIN OUTCOME MEASURES:The NSCs and its differential potential were identified by surface marker nestin and immunocytochemistry.RESULTS:The neurospheres formed after 1 week culture,and grew with time prolonged.The culture system was changed into adherent culture at 4th week.At 24 hours after plated on the poly-omithine/fibronectin pre-coated dish,neurospheras adhered to the dish and many cells migrated out of neurnshperes and proliferated as monolayer.The immunocytochemistry showed that NSCs which culture medium contain EGF and bFGF preserve neatin positive,and the cmNSCs that without EGF and bFGF differentiated into astrocytes (GFAP positive),neurons (Tuj-1 positive) and oligodendrocytes (O4 positive).CONCLUSION:The culture system which combines suspension culture and adherent culture is fit for the culture of cmNSCs.The cultured cmNSCs has potential to differentiate into astrocytes,neuronal cells and oligodendrocytes.

Atherosclerosis ; diagnosis ; Humans ; Tomography, Optical Coherence ; methods

Objective: To construct the extracellular region of PCP-2(PCP-2EC) and the immunoglobin IgG Fc fusi on protein expression vector,and then express and purify the soluble PCP-2EC/Fc fusion protein for the study of its function in neuronal adhesion. Methods: PCP-2 extracellular region was amplified and cloned into an expression vector pIGplus containing human IgG Fc; PCP-2EC/Fc fusion protein was expressed by COS-7 and 293 cells transfected by the constructed plasmid and purified by protein A. The purified fusion protein was used as substrate to study its function in neuronal adhesion. Results: PCP2 extracellular region was cloned into IgG Fc expression vector successfully; PCP 2EC/Fc fusion protein was expressed and purified in mammal cells; and the purified fusion protein promoted neuronal adhesion. Conclusion:PCP 2EC/Fc fusion protein expression system is successfully constructed and the purified fusion protein can promote neuronal adhesion. These results lay a foundation for the research on the PCP-2 function in neuronal adhesion and the further functional study in the nervous system and other fields.

Objective:To analyze the significance of equipment custodian routine maintenance work and then offer some suggestion of countermeasure.Methods: Twenty four cases of maintenance cassette sterilizer were divided into test group(12 cases), equipment custodian had been trained; control group (12 cases)has not received training. In addition, we analyzed separately the maintenance work with the risk theory and cost-benefit.Results: The total number of failures in the test group was 13, obviously lower than 32 in the control. The risk of all devices under control, the maintenance of the benefits far outweigh the cost.Conclusion:Training equipment custodian, can reduce the incidence of failure, are worthy of promotion.

Currently campus violence including the murder case in which college students shot teachers has shocked the whole society,arousing various issues for educators.This paper holds that the carrying out of life education is one key solution to those new rising social issues.Through discussing the origin of students' ignorance of human lives,the significance and measures of performing life education are explored in this paper.

Objective: To construct the extracellular region of PCP-2(PCP-2EC) and the immunoglobin IgG Fc fusi on protein expression vector,and then express and purify the soluble PCP-2EC/Fc fusion protein for the study of its function in neuronal adhesion.Methods: PCP-2 extracellular region was amplified and cloned into an expression vector pIGplus containing human IgG Fc; PCP-2EC/Fc fusion protein was expressed by COS-7 and 293 cells transfected by the constructed plasmid and purified by protein A.The purified fusion protein was used as substrate to study its function in neuronal adhesion.Results:PCP-2 extracellular region was cloned into IgG Fc expression vector successfully; PCP-2EC/Fc fusion protein was expressed and purified in mammal cells; and the purified fusion protein promoted neuronal adhesion.Conclusion:PCP-2EC/Fc fusion protein expression system is successfully constructed and the purified fusion protein can promote neuronal adhesion.These results lay a foundation for the research on the PCP-2 function in neuronal adhesion and the further functional study in the nervous system and other fields.

Objective To investigate the protective effects of isovolemic hemodilution with 6% hetastarch (HAES) and ligustrazine on myocardium against ischemia/reperfusion injury. Methods Thirty-two healthy male rabbits were anesthetized with 3% pentobarbital 30 mg?kg-1 , tracheotomized and mechanically ventilated. Left femoral artery and vein were cannulated for direct BP monitoring, blood collection and fluid infusion. Chest was opened and myocardial ischemia was produced by temporary ligation of the anterior descending branch of left coronary artery. Myocardial ischemia was confirmed by elevation or depression of S-T segment and /or high T-wave. The animals were randomly divided into 4 equal groups with 8 animals in each group: Ⅰ control group was subjected to 45 min myocardial ischemia followed by 180 min reperfusion without any treatment; Ⅱ hemodilution group in which 9 ml Ⅲ kg-1 blood was removed and blood volume was maintained by simultaneous infusion of equal volume of 6% HAES at 20 min after myocardial ischemia was started; Ⅳ ligustrazine group received ligustrazine injectio 20 mg?kg-1 iv 20 min before and 40 min after myocardial ischemia was started; Ⅱ hemodilution + ligustrazine group in which hemodilution was performed as in group Ⅲ and ligustrazine injectio was given iv as in group Ⅲ . BP and HR were recorded before during and at the end of myocardial ischemia, and at 30, 60, 120, 180 min of reperfusion. Hct was measured before and after hemodilution. Blood samples were taken for determination of plasma CPK and LDH activities before ischemia (T0), at the end of 45 min ischemia (T1) and at the end of 180 min reperfusion (T2 ) . At the end of the experiment, myocardial tissue 0.2 g was obtained from ischemic and non-ischemic area for determination of myocardial CPK and LDH activities and election microscopic examination. Results (1) In all four groups plasma CPK and LDH activities were significantly increased after ischemia (T1 ) and were increased further after reperfusion (T2) (P

Objective To identify the charactor of wide QRS complex tachycardia( WCT)throuGh transesophaGeal atrial pacinG( TEAP ). Methods TEAP and intracadiac electrophysioloGical examination infoamation of l2 cases WCT were collected and analyzed from January to February in 20l2 of Wuhan Asia Heart Hospital. Results Comparison of TEAP and intracadiac electrophysioloGical examination showed that l0 in l2 patients were match. Conclusion TEAP is a rapid and convenient method to diaGnose most WCT.