Objective To construct a shuttle plasmid for inducible gene expression in Borrelia burgdorferi (B.burgdorferi) with an advantage of flexible genetic manipulation.Methods The IPTG-inducible lac repressor/operator system from Escherichia coli (E.coli) was adopted and modified in the current study.The plasmid shuttle vector was developed by inserting multiple cloning sites,FLAG and HA tags into the shuttle vector by molecular cloning approaches.The target gene was inserted at the site under the control of the promoter (Tn5 derivate) in plasmid pQE30.This promoter contained two lac operators and a codonoptimized lacI gene driven by flaB promoter.Results A plasmid shuttle vector,pJJ275,was successfully constructed with the ability to express target genes in B.burgdorferi in the presence of IPTG.By using this system,a HA-tagged rpoS gene was introduced into the typical infectious strain B.burgdorferi B31.The target gene expression induced by IPTG was confirmed at transcriptional and translational levels.The RpoS dependent virulence factor of Borrelia,OspC,was also detected,indicating that the expressed protein was functional.Conclusion The constructed plasmid shuttle vector can express exogenous genes in B.burgdorferi with an inducible feature and an advantage of flexible genetic manipulation.It can be applied for genetic manipulation of B.burgdorferi involved in gene regulation and complementation.

Objective:To investigate the expression of Rho GTPases signaling pathway in non-small cell lung carcinoma (NSCLC)and its clinical significance.Methods:Molecules of Rho GTPases signaling pathway including RhoC, E-cadherin, MMP-2, and MMP-9 were detected by RT-PCR and immunohistochemistry in 36 specimens of non-small cell lung carcinoma (NSCLC). The relationship between RhoC mRNA and prognosis of patients was evaluated by using Kaplan-Meier survival curve analysis.Results:There was a significant difference in the expression of RhoC mRNA between NSCLC tissues and para-cancerous tissues (P<0.01). The expression of RhoC mRNA in NSCLC was not correlated with gender, age, invasion degree, lymph node metastasis, tumor size, histological classification, and differentiation degree but correlated with different TNM stage (P<0.05). There was a significant positive correlation between the expression of RhoC protein and MMP-2 protein (r=0.474, P=0.003). The survival time of patients with weak expression of RhoC mRNA was longer than those with over-expression of RhoC mRNA, but there was no significant difference between the two groups (P>0.05). Conclusion:Over-expression of RhoC mRNA is closely correlated with the tumorigenesis and progression of NSCLC, and may be related with invasion and metastasis of NSCLC early to middle stage.

Objective To compare the diagnostic efficiency of the thyroid imaging reporting and data system (TIRADS), the Bethesda system for reporting thyroid cytopathology(BSRTC) and BRAFV600E detection, and their combined use in the differentiation between benign and malignant thyroid nodules. Methods One hundred and twenty eight patients with 128 thyroid nodules who were scheduled for ultrasound-guided fine-needle aspiration biopsy (FNAB) were recruited for the study. All of them underwent ultrasound, fine-needle aspiration cytology(FNAC) examination, and BRAFV600E detection. TIRADS and BSRTC systems were adopted to judge the ultrasound and FNAC results. The receiver operating characteristic curve was established to assess the diagnostic value of each method. Results The sensitivity, specificity, and AUC of TIRADS were 74. 3%, 84. 5%, and 0. 794, respectively. BSRTC had higher specificity(98. 3%) and equal sensitivity compared to TIRADS. The sensitivity, specificity, and AUC of BRAFV600E detection were the highest ones among the three methods. Combinations of different methods could increase the diagnostic sensitivity and accuracy. The combination of FNAC and BRAFV600E detection significantly increased the diagnostic efficiency(AUC=0. 984), with sensitivity 98. 6% and specificity 98. 3%. Conclusions The diagnostic value of BRAFV600E detection in the differentiation of benign and malignant thyroid nodules is better than both TIRADS and BSRTC, and the combination of FNAC and BRAFV600E detection reaches the best diagnostic efficiency.