ObjectiveTo observe the effect of selective posterior rhizotomy (SPR) on children with spastic cerebral palsy.Methods517 spastic cerebral palsy cases were operated on by SPR, and a following up was performed for 24 months. After operation, curative effect of SPR was examined and evaluated.Results298 cases had excellent effect (57.6%); 187 cases had good effect (36.2%).Conclusion SPR is very effective for children with spastic cerebral palsy.

Objective To evaluate laparoscopic balloon nasobiliary biliary drainage (LBNBD),vs ureteral catheter drainage in one stage laparoscopy,choledochoscopy and duodenoscopy choledocholithotomy and primary closure of the small calibered common bile duct (diameter 0.3-0.8 cm).Methods During the period of Apr 2010 to Nov 2016 102 cases were enrolled including 50 cases receiving LBNBD and 52 cases using ureteral catheter drainage.Results Between the two groups,LBNBD was superior to ureteral catheter drainage in all the following parameters:the operation time,intraoperative blood loss,postoperative liver function,blood amylase and other laboratory indicators,gastrointestinal function recovery time,gastrointestinal symptoms and electrolyte imbalance,postoperative hospital stay,and bile duct drainage time with all differences statistically significant (P ＜ 0.05).Bile drainage differences during the postoperative first 3 days (averagely 200-400 ml a day) were not statistically different (P ＞ 0.05).Postoperative pancreatitis,bile leakage,and hemobilia were not statistically different (P ＞ 0.05).Conclusions The use of LBNBD is safe and effective in endoscopic choledocholithotomy in cases of small calibered common bile duct.

Overlap extension PCR is a common method for site-directed mutagenesis. As objective gene sequence growing longer, it is often difficult to obtain the target product in the second round of PCR, and it is highly possible to introduce unexpected mutations into a long gene fragment by PCR. To circumvent these problems, we can only amplify a small gene fragment which contain the target mutation by overlap extension PCR, and then ligate it with vector to get target plasmid. If the restriction site at the end of the amplified fragment was not a single one on plasmid vector, double fragments ligation method could be used to construct target plasmid. Partial amplification, combined with double fragments ligation, could solve lots of problems in long gene mutagenesis. Taking retinoblastoma gene 1 S780E mutagenesis as an example, it is difficult to amplify whole retinoblastoma gene 1 by overlap extension PCR because of long fragment interfering the overlapping extension of second round PCR. However, it is relatively easy to amplify the F3 (1 968-2 787) fragment which contains target mutation S780E. There is a Nhe I site which can be used for ligation on 5' end of F3 fragment, but another Nhe I site on the plasmid restrained from doing so directly. In order to circumvent this obstacle, we ligated F3 fragment, combining with F2 (900-1 968) fragment which was digested from wild type plasmid, with the vector which contain F1 (1-900) fragment of the gene. That double fragments ligated with one vector at the same time, though less efficient, can recombine into a complete plasmid. The sequences of the two selected recombinant plasmids were consistent with the target mutation, which verified the feasibility of this scheme. As an improvement of overlap extension PCR, partial amplification and double fragments ligation methods could provide solutions for site directed mutagenesis of many long genes.
Base Sequence ; Cloning, Molecular ; Genetic Vectors ; genetics ; Mutagenesis, Site-Directed ; methods ; Nucleic Acid Amplification Techniques ; Plasmids ; Polymerase Chain Reaction

OBJECTIVE： To determine and compare the contents of catalpol and aucubin in different parts （root， stem， leaf and flower） of wild Centranthera grandiflora， and to provide reference for the selection of medicinal parts and source development. METHODS： HPLC method was used to determine the contents of catalpol and aucubin in root， stem， leaf and flower of wild C. grandiflora， and the contents of different parts were analyzed comparatively. The determination of catalpol was performed on Agilent TC-C18 column with mobile phase consisted of methanol-0.1％ phosphoric acid （1 ∶ 99， V/V） at the flow rate of 1 mL/min； the detection wavelength was set at 210 nm， and sample size was 20 μL. The column temperature was 35 ℃； the determination of aucubin was performed on SPHERI-5RP-C18 column with mobile phase consisted of acetonitrile-water （3 ∶ 97， V/V） at the flow rate of 1 mL/min； the detection wavelength was set at 205 nm， and sample size was 20 μL； the column temperature was 25 ℃. RESULTS： The linear range of catalpol and aucubin were 0.061 5-3.321 and 0.000 36-0.216 mg/mL （all r＝0.999 9）. The limits of detection were 0.016 and 0.007 μg/mL. The limits of quantitation were 0.052 and 0.023 μg/mL. RSDs of precision， stability （24 h） and reproducibility tests were all lower than 2.00％ （n＝6）. The average recoveries were 99.34％ and 99.61％， and RSDs were 1.06％ and 1.12％， respectively （n＝6）. The average content of catalpol in root， stem， leaf and flower wild C. grandiflora were 1.609， 3.030， 11.095 and 1.921 mg/g， respectively. The contents of aucubin in different parts were 0.441， 0.020， 0.005 and 0.006 mg/g，respectively. CONCLUSIONS：The established HPLC method meets the requirements of quantitative analysis. Catalpol is mainly distributed in the leaves of wild C. grandiflora， and aucubin is mainly distributed in the roots of wild C. grandiflora. The experimental conclusion provides a reference for the reasonable selection of different medicinal parts as raw materials to develop medicine with different efficacy.