Objective:To construct the eukaryotic expression plasmid of EGFP/H2B and express it in the Human embryo kidney cells(HEK293).Methods:Part H2B gene was obtained by RT-PCR from cultured HEK293 cell's whole RNA.The sequence encoding H2B was cloned into pEGFP-C1 to construct the eukaryotic expression vector.After confirmed by enzymatic digestion and sequencing,the recombination plasmid was transfected to HEK293 cells by lipofectamine technology for observation of the fusion protein's expression.Results:The recombination plasmid of pEGFP/H2B was constructed successfully.Compared with the dispersing green fluorescence among the whole cells in the control cells transfected with the mock plasmid(pEGFP-C1),the green fluorescence was concentrated in the nuclei of HEK293 cells transfected with pEGFP/H2B and distributed with the chromosomes in the mitotic cells.Conclusions:The successful construction of pEGFP/H2B recombination plasmids establishes an available survey system to study the chromosome segregation mechanism.

Objective To investigate the diagnostic value of improved pleural biopsy combined with biomarker and cytology detection in pleural effusion of unknown origin.Methods The clinical data in 216 cases of pleural effusion were respectively analyzed including 106 cases of tuberculous pleural effusion(tuberculosis group) and 110 cases of malignant pleural effusion(malignant group).All cases were performed the improved pleural biopsy,cytology examination and detection of pleural effusion ADA,CEA and LDH,and serum CEA.The pleural biopsy diagnosis rate was performed the statistics,and pleural effusion ADA,CEA and LDH,serum CEA,and pleural effusion CEA/serum CEA were compared between the two groups.Results Among 216 cases,241 times of pleural biopsy puncture were conducted,the first time puncture success rate was 94.9 ％ (205/216).Having the diagnostic value among pathological results of pleural biopsy materials in first puncture success accounted for 58.8％ (127/216),and the overall diagnosis rate was 65.3 ％ (141/216).The incidence rate of adverse reactions was 5.8 ％ (14/241).In the tuberculosis group,no case showed cytology tumor cell positive,while the cytology tumor cell positive rate in the malignant group was 54.5％ (60/110);pleural effusion CEA and LDH,serum CEA and pleural effusion CEA / serum CEA levels and positive rates in the malignant group were significantly higher than those in the tuberculosis group,while the pleural effusion ADA level and positive rate were significantly lower than those in the tuberculosis group the difference was statistically significant(P＜0.01).Conclusion Improved pleural biopsy,pleural effusion cytology,pleural effusion biomarkers have a certain limitation in alone auxiliary diagnosis of pleural effusion.The various indicators can be combined to determine the etiology of pleural effusion in clinic for guiding treatment.

Objective To explore the mechanism of hBUB1 gene in the developing of spontaneous abortion embryos with numerical chromosomal abnormalitv.Methods Quantitative real-time RT-PCR and Western blot were used to determine the mRNA and protein level of hsMAD2 gene both in spontaneous abortion embryos with numerical chromosomal abnormality(experimental group)and with numerical chromosomal normality(control group).Recombinant shRNA plasmids targeting hBUB1 gene was constructed to inhibit the expression of endogenous hBUB1 genes in embryonic cells.Interference efficiency was demonstrated by fluorescent quantitative PCR and Western blot.The inhibitory rate of cell proliferation was measured by MTT assay and cells-cycle was assessed by flow cytometry.Resuns Western blot analysis showed that protein level of hBUB1 in the experimental group was decreased signifieandv(the rate of positivity and strong positivity were 8%and 93.5%,respectively,P＜0.05)compared with the control group.The expression of hBUB1 gene in embryonic cells was significantly and specially inhibited by shRNA plasmids (the mRNA level before and after treatment witll RNAi were 0.196±0.067 and 0.042±0.006,respectively,P＜0.05).The inhibitory rate of cell proliferation was increased to 62%from 4%at 48 h after transfection.The rate of G2/M phase cells was decreased after transfection with efficient shRNA(control group:40.2%and 41.3%,test group:21.3%).Conclusions Down-regulation of hBUB1 gene leads to the inhibition of cell proliferation and the arrest of cell-cycle.It also probably plays an important role in the development of spontaneous abortion embryos with numerical chromosomal abnormality.The clinical relevance warrant further investigation.

Objective:To investigate the value of serum programmed cell death molecule 5 (PDCD5) protein expression in early prediction of gastric cancer and its clinical significance.Methods:A total of 103 patients with gastric cancer who were treated in Yuechi County People′s Hospital in Sichuan Province from March 2014 to March 2016 and 80 healthy people who underwent physical examinations (control group) in the same period were selected as subjects. The serum level of PDCD5 protein were detected by enzyme-linked immunosorbent assay. The diagnostic performance of serum PDCD5 protein on gastric cancer was evaluated by receiver operating characteristic curve. The patients with gastric cancer were divided into low-level group (50 cases) and high-level group (53 cases) according to serum PDCD5 protein level. The relationship between serum PDCD5 protein level and clinical data in patients with gastric cancer was analyzed by χ2 test. Univariate and multivariate Cox regression models were used to analyze independent risk factors for survival and prognosis of gastric cancer. Kaplan-Meier method was used to map survival curves of gastric cancer patients with different levels of serum PDCD5 protein. Results:Serum PDCD5 protein level in gastric cancer group was significantly lower than that in control group: (0.82 ± 0.30) mg/L vs. (1.26 ± 0.39) mg/L, and there was statistical difference ( t=8.628, P<0.01). Serum PDCD5 protein level in patients with gastric cancer was related to tumor TNM stage and tumor invasion ( P<0.05), but not related to gender, age, body mass index (BMI), tumor size, lymph node metastasis, tumor type and tumor differentiation ( P<0.05). The area under curve (AUC) of serum PDCD5 protein in the diagnosis of gastric cancer was 0.810 (95% CI 0.747 to 0.873), with a sensitivity of 71.8%, and a specificity of 76.3% ( Z=9.641, P<0.01). Serum PDCD5 protein level was an independent risk factor for poor prognosis in patients with gastric cancer ( P<0.05). The 5-year survival rate in low-level group was significantly lower than that in high-level group: 32.0% vs. 62.3%, and there was statistical difference ( χ2=18.422, P<0.01). Conclusions:The serum PDCD5 protein level in patients with gastric cancer is significantly decreased. Low serum PDCD5 protein level is independent risk factors for poor prognosis of patients with gastric cancer.