Objective For the purpose of exploring the influence exerued by the amount of template DNA on the results of automatic fluerescent detection. Method For the purpose of buiding the optimal amount of template DNA amplified, we have analyzed different amount of template DNA (9947A) by 3100 Genetic Analyzer with the multiplexing Profiler Plus Kit. Results The optimal amounts of template DNA amplified were between 0.31ng and 2. 5ng by using 3100 Genetic Analyzer. Conclusion It will produce the incorrect analytic results if the amounts of template DNA were too high or too low.

imaging features of myositis ossificans have some characteristics. Misdiaguosis could be avoided when the disease was evaluated with the course.

Child, Preschool ; Fever ; etiology ; Histoplasmosis ; complications ; diagnosis ; pathology ; Humans ; Jaundice ; etiology ; Male

Objective:To investigate the effect of miRNA-5089-5p (miR-5089-5p) on proliferation and migration ability of esophageal cancer in vitro and its relationship with the expression of cathepsin B (CTSB) gene.Methods:Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression levels of miR-5089-5p in 31 tissue samples from patients who underwent esophageal cancer resection and the corresponding pericarcinomatous tissues between in March 2017 and in December 2019 at Huangshi Central Hospital of Edong Healthcare Group, and TE-13, EC9706, Eca109, KYSE30 cell lines and normal esophageal mucosal epithelial HET-1A cells. The esophageal cancer cells with the lowest expression level of miR-5089-5p were divided into 2 groups: miR-5089-5p group transfected with miR-5089-5p mimics and the negative control group with negative control sequence. qRT-PCR was used to detect the expression level of miR-5089-5p after transfection for 48 h. CCK-8 method and scratch healing test were used to detect the proliferation and migration ability of cells in the two groups. The online tools microRNA.org and Deepbase v2.0 were used to predict the target genes of miR-5089-5p. The dual luciferase reporter gene assay was used to verify the target gene of miR-5089-5p. qRT-PCR and Western blot were used to detect the expression level of target genes in the two groups. The expressions of cell proliferation-related protein (PCNA and Ki-67) and migration-related protein (N-Cadherin and Twist) were detected by using Western blot.Results:The relative expression level of miR-5089-5p in esophageal cancer and pericarcinomatous tissues was 1.54±0.53 and 7.07±1.25, respectively ( t = 24.06, P < 0.01). The relative expression level of miR-5089-5p in the esophageal cancer cell lines was lower than that of normal esophageal mucosal epithelial HET-1A cells (all P < 0.05), and the cell line with the lowest relative expression was Eca109 cells (0.12±0.03). Compared with the negative control group, the proliferation ability of Eca109 cells in miR-5089-5p group was gradually reduced with the transfection time extension, and the difference was statistically significant between the two groups since 48 h (all P < 0.05), and the migration ability was also reduced [scratch healing rate: (29±5)% vs.(64±8)%, t=3.91, P < 0.01]. The online tool predicted that the target gene of miR-5089-5p might be CTSB, and the dual luciferase reporter gene assay confirmed that miR-5089-5p complemented CTSB 3'UTR. qRT-PCR results showed that compared with the negative control group, the relative expression level of CTSB mRNA in Eca109 cells of miR-5089-5p group was reduced (0.23±0.04 vs.1.01±0.09, t = 8.27, P < 0.01). Western blot results showed that the expression level of CTSB protein was reduced, and the expression levels of cell proliferation-related protein PCNA, Ki-67 and cell migration-related protein N-Cadherin, Twist were also reduced. Conclusions:The expression level of miR-5089-5p in esophageal cancer tissues and cell lines is low. miR-5089-5p can inhibit proliferation and migration of esophageal cancer Eca109 cells. The mechanism may be achieved by down-regulating CTSB gene expression.

Vitexin, a naturally occurring flavone glycoside in plants, has many pharmacological effects, which is widely distributed in nature. This paper reviewed the research progress of the distribution of vitexin in the plant resources and its pharmacological effects, and summarized its application prospects, aiming to provide a useful reference for the development of vitexin-enriched plant resources.
Animals ; Antineoplastic Agents ; pharmacology ; Antioxidants ; pharmacology ; Apigenin ; pharmacology ; Humans ; Hypoglycemic Agents ; pharmacology ; Myocardial Infarction ; drug therapy ; Plant Dispersal

Anion Exchange Protein 1, Erythrocyte ; metabolism ; CD56 Antigen ; metabolism ; Desmin ; metabolism ; Desmoplastic Small Round Cell Tumor ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Humans ; Lymphoma ; metabolism ; pathology ; Male ; Middle Aged ; Rhabdomyosarcoma ; metabolism ; pathology ; Sarcoma, Ewing ; metabolism ; pathology ; Testicular Neoplasms ; metabolism ; pathology ; surgery ; Vimentin ; metabolism

Gene knockout by ZFNs (zinc-finger nucleases) is efficient and specific, and successfully applied in more than 10 organisms. Currently, it is unclear whether this technology can be used for knocking-out enhanced green fluorescent protein (EGFP) gene in transgenic goats. Here we constructed and used ZFN-coding plasmids to produce genetic knockouts in the cells of cloned fetus produced from donor cells by microinjection of EGFP gene. Following introduced plasmids into caprine primary cultured fetus fibroblasts by electroporation, targeting of a transgene resulted in sequence mutation. Using the flow cytometric analysis, we confirmed the disappearance of EGFP expression in treated cells. Sequence from PCR products corresponding to targeted site showed that insertion of a G into the exon of EGFP resulted in frame shift mutation. These results suggest that ZFN-mediated gene targeting can apply to caprine fetus fibroblasts, which may open a unique avenue toward the creation of gene knockout goats combining with somatic cell nuclear transfer.
Animals ; Base Sequence ; Cloning, Organism ; Electrophoresis ; Endonucleases ; genetics ; metabolism ; Fetus ; Fibroblasts ; metabolism ; Gene Knockout Techniques ; Gene Targeting ; methods ; Goats ; Green Fluorescent Proteins ; genetics ; Molecular Sequence Data ; Mutation ; Zinc Fingers

Objective To investigate the dynamic expressions of exchange protein directly activated by cyclic adenosine monophosphate (cAMP) (Epac) in rat model of hepatic fibrosis(HF).Methods Forty-two male SD rats were divided into control group (n = 6) and model group (n = 36)which was divided into six subgroups of day 4, week 1, week 2, week 4,week 6 and week 8 with six rats in each subgroup. The rat model of HF was established by intraperitoneal injection of dimethylnitrosamine (DMN). The pathological changes of liver were observed by Hematoxylin-Eosin and Masson staining. Reverse transcription-polymerase chain reaction (RT-PCR),immunohistochemistry and Western blot were employed to detect the mRNA and protein expressions of Epac1, Epac2 and transforming gronth factor (TGF)β1 during the process of modeling and localization in the liver. The statistical analysis was done using one-factor ANOVA, LSD-t test,Dunnett T3 test and Pearson linear correlation analysis. Results Rat model of liver fibrosis was established successfully. In control group, Epac1 (0. 031 28±0. 008 96) and Epac2 protein (0.034 43±0. 002 45) mainly expressed in the cytoplasm of hepatocytes. In model group, the level of Epac1 decreased at day 4 (0. 023 97±0. 003 81) and week 1 (0. 015 81±0. 002 48) ,then began to increase at week 2 of modeling and peaked at week 6 (0. 039 54±0. 001 43), which had statistical significance compared to the control group (t= 5.47,11.58 and - 6.18, respectively; all P＜0.05). Epac2 protein expression declined after modeling, reached the lowest level at week 4 (0. 011 21 ±0. 001 32), which had statistical significance compared to the control group (t= 24. 50, P＜0. 05). TGFβ1 protein expression increased after modeling and peaked at week 4 (0. 011 30±0.001 03) which had statistical significance (t= -23. 36, P＜0. 05) compared to the control group (0. 002 08 ±0. 000 18). The expressions of Epac1, Epac2 and TGFβ1 mRNA were consistent with the trend of protein levels.Correlation analysis showed that Epac1 protein was positively correlated with the course of HF (r =0. 703, P＜0.01 ), while Epac2 protein was negatively correlated (r = - 0. 409, P＜0.05). Conclusions During the progression of HF, Epac1 expression tends to decrease firstly and increase afterwards,while Epac2 expression declines continually. Epac may be involved in the pathogenesis of HF.

Objective To use the fluorescence PCR‐melting curve method to detect CYP2C9 and VKORC1 gene polymorphism in Xinjiang Hui population ,to analyze their gene distribution and gene mutation frequency ,and to evaluate the clinical applicability of the fluorescence PCR‐melting curve method .Methods The fluorescence PCR‐melting curve method and sequencing method were adopted to contrastively detect CYP2C9*2 ,CYP2C9*3 and VKORC1(‐1639G/A)gene polymorphism .Results Among detected 228 Xinjiang Hui individuals ,199 cases of CYP2C9*1/*1 ,2 cases of CYP2C9*1/*2 ,26 cases of CYP2C9*1/*3 and only 1 case of CYP2C9*3/*3 were detected ,no case of CYP2C9*2/*2 and CYP2C9*2/*3 was detected .Two kinds of allele G and A were detected for VKORC1(‐1639G/A) ,in which VKORC1‐1639G/G type was detected in 2 cases ,VKORC1‐1639G/A type was detected in 39 cases and VKORC1‐1639A/A type was detected in187 cases ,compared with the sequencing method ,the results of the fluorescence PCR‐melting curve method were completely consistent .Conclusion Xinjiang Hui population also has CYP2C9 gene *2 ,*3 loci and VKORC1 gene(‐1639G/A) locus polymorphism ,their occurrence frequency has a certain difference with Xingjiang Uygur and other regional populations ,the adopted fluorescence PCR‐melting curve method used in the gene polymorphism detection can meet clinical detection requirements .

Objective To explore the clinical value of guide-wire shaping in subclavian vein catheter-ization.Methods Totally 400 patients requiring right subclavian vein catheterization were equally divided into two groups according to the clinic date: intervention group ( with guide-wire shaping , n =200 ) and control group (without guide-wire shaping, n=200).The catheterization was carried out by the same doctor .The rates of ectopic wire were compared between the two groups .Results The overall success rate of catheteriza-tion was 98.25%(393/400) [98.5% (197/200) in intervention group and 98.0% (196/200) in control group, P=0.500].The incidence of catheter displacement was 1.02%(2/197) in intervention group, which was significantly lower than that [7.14% (14/196)] in control group (P=0.002).Conclusion As a sim-ple procedure , guide-wire shaping can effectively prevent catheter displacement during catheterization .