Objective To construct pIRES2-ZsGreen1/FⅨ expression vector,using the pcDNA/ FⅨ plasmid containing FⅨ cDNA as template,and express in HEK-293 cells.Methods The total ORF of F Ⅸ gene was amplified from pcDNA/F Ⅸ plasmid,then the amplified fragment was clonded into the pIRES2-ZsGreen1 vector using the Infusion enzyme.The positive clones of eukaryotic expression vector of pIRES2-ZsGreen1/F Ⅸ were screened and expanded after transfection,then were constructed and confirmed by PCR and sequencing.Transient expression experiments were performed using HEK-293 cells transfected with the expression vectors and observed the expression of ZsGreenl protein by confocal laser microscope.The relative expression levels of F Ⅸ mRNA,protein and F Ⅸ activity (F Ⅸ ∶ C) were detected by real time PCR (RT-PCR),immunofluorescence microscopy,One-Stage method,respectively.Results The expression vector,pIRES2-ZsGreen1/F Ⅸ,was successfully constructed and expressed in HEK-293 cells.RT-PCR detected the expression of F Ⅸ mRNA in HEK-293 cells and the immunofluorescence microscopy showed F Ⅸ protein distributed in the surrounding of nucleus.F Ⅸ ∶ C of cell lysates and cell culture fluid transfected with the expression vectors were (92.03 ± 0.29)％ and (86.89 ± 8.78)％,respectively;while both F Ⅸ ∶ C of cell lysates and cell culture fluid transfected with or without the expression vectors were 0.Conclusion The experimental results showed the expression vector,pIRES2-ZsGreen1/F Ⅸ,was successfully constructed,which provided experiment basement for the follow study on the location,function and molecular pathology of hemophilia B.

Objective To analyze the risk factors of intraoperative hypothermia in patients un-dergoing radical thyroidectomy under laparoscopy and to construct a nomogram prediction model.Methods A total of 336 patients who underwent laparoscopic radical thyroidectomy were selected as study subjects.According to intraoperative body temperature,they were divided into hypothermia group(195 cases)and normal temperature group(141 cases).The risk factors of intraoperative hy-pothermia in patients undergoing laparoscopic radical thyroidectomy were analyzed using the Logistic regression model.The nomogram prediction model was constructed using R software.The prediction performance of the nomogram prediction model was evaluated using the receiver operating characteris-tic(ROC)curve,calibration curve,and Hosmer-Lemeshow goodness-of-fit test.Results There were significant differences in age,intraoperative blood loss,amount of fluid infusion,and operation time between the two groups(P＜0.05).Multivariate Logistic regression analysis showed that age＞60 years,intraoperative blood loss＞60 mL,amount of fluid infusion＞1 000 mL,and operation time＞2.5 h were risk factors for intraoperative hypothermia in patients undergoing laparoscopic radical thy-roidectomy(P＜0.05).A nomogram prediction model was constructed based on the four risk factors.The ROC curve showed that the area under the curve was 0.912(95％CI,0.884 to 0.941).The calibration curve and Hosmer-Lemeshow goodness-of-fit test showed that the slope of the calibration curve was close to 1(x2=9.140,P=0.243).Conclusion The nomogram prediction model based on age,intraoperative blood loss,amount of fluid infusion,and operation time has good predictive value for intraoperative hypothermia in patients undergoing laparoscopic radical thyroidectomy.

Objective To analyze the risk factors of intraoperative hypothermia in patients un-dergoing radical thyroidectomy under laparoscopy and to construct a nomogram prediction model.Methods A total of 336 patients who underwent laparoscopic radical thyroidectomy were selected as study subjects.According to intraoperative body temperature,they were divided into hypothermia group(195 cases)and normal temperature group(141 cases).The risk factors of intraoperative hy-pothermia in patients undergoing laparoscopic radical thyroidectomy were analyzed using the Logistic regression model.The nomogram prediction model was constructed using R software.The prediction performance of the nomogram prediction model was evaluated using the receiver operating characteris-tic(ROC)curve,calibration curve,and Hosmer-Lemeshow goodness-of-fit test.Results There were significant differences in age,intraoperative blood loss,amount of fluid infusion,and operation time between the two groups(P＜0.05).Multivariate Logistic regression analysis showed that age＞60 years,intraoperative blood loss＞60 mL,amount of fluid infusion＞1 000 mL,and operation time＞2.5 h were risk factors for intraoperative hypothermia in patients undergoing laparoscopic radical thy-roidectomy(P＜0.05).A nomogram prediction model was constructed based on the four risk factors.The ROC curve showed that the area under the curve was 0.912(95％CI,0.884 to 0.941).The calibration curve and Hosmer-Lemeshow goodness-of-fit test showed that the slope of the calibration curve was close to 1(x2=9.140,P=0.243).Conclusion The nomogram prediction model based on age,intraoperative blood loss,amount of fluid infusion,and operation time has good predictive value for intraoperative hypothermia in patients undergoing laparoscopic radical thyroidectomy.

Adult ; Aged ; Autoimmune Diseases ; genetics ; Case-Control Studies ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; Receptors, IgG ; genetics ; Thrombocytopenia ; genetics ; Young Adult

Computational Biology ; Factor VII ; Factor VII Deficiency ; Genetic Testing ; Humans ; Pedigree