Targeting induced local lesions in genomes (TILLING) is a reverse genetics method for functional genomics research.It is possible to screen for point mutations in the populations of EMS mutagenesis with highthroughput and lowcost. EcoTILLING a method based on TILLING ,was developed for detecting multiple types of polymorphisms in germplasm collections,such as single nucleotide polymorphism,small deletion and insertion etc.Rice is a very important food crop and a model plant for genome research also. There are complete genome sequence and a lot of other bioinformatics resources about it.So the markerassisted breeding is becoming more and more important in rice breeding. Some issues based on TILLING about identifying germplasm based on gene sequence,EMS mutagenesis breeding,developing functional marker in rice breeding in future were discussed.

Aged ; Aged, 80 and over ; Anti-Bacterial Agents ; pharmacology ; Coal Mining ; Cross Infection ; complications ; microbiology ; Drug Therapy, Combination ; therapeutic use ; Humans ; Male ; Methicillin ; pharmacology ; Methicillin Resistance ; Microbial Sensitivity Tests ; Middle Aged ; Pneumonia ; drug therapy ; etiology ; Silicosis ; classification ; complications ; Staphylococcus aureus ; drug effects ; Treatment Outcome

This study was aimed to construct a pEGFP-N1 vector of von Willebrand factor cleaving protease (ADAMTS13, a disintegrin and metalloprotease with a thrombospondin type 1 motifs 13) so as to pave the way for further studying its synthesis and secretion. Human full-length cDNA sequence of ADAMTS13 was acquired by polymerase chain reaction (PCR) with Phusion(®) High-Fidelity (NEB), then the PCR product was double digested with EcoRI and XhoI. After digestion, the ADAMTS13 cDNA sequence was purified and recombined with the pEGFP-N1 vector. The DNA sequence analysis showed that ADAMTS13 was ligated to the pEGFP-N1 vector correctly. After transient expression in HeLa cells, the expression of EGFP could be detected by fluorescent microscopy, and the expression of ADAMTS13 protein could be detected by SDS-PAGE and Western blot. It is concluded that the ADAMTS13-pEGFP-N1 vector is successfully constructed, and it can be widely used in further research on the mechanism of the synthesis and secretion of ADAMTS13.
ADAM Proteins ; genetics ; ADAMTS13 Protein ; Gene Expression ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; HeLa Cells ; Humans ; Transfection

Citri Fructus identification by fingerprint and chemometrics was investigated in this paper. Twenty-three Citri Fructus samples were collected which referred to two varieties as Cirtus wilsonii and C. medica recorded in Chinese Pharmacopoeia. HPLC chromatograms were obtained. The components were partly identified by reference substances, and then common pattern was established for chemometrics analysis. Similarity analysis, principal component analysis (PCA) , partial least squares-discriminant analysis (PLS-DA) and hierarchical cluster analysis heatmap were applied. The results indicated that C. wilsonii and C. medica could be ideally classified with common pattern contained twenty-five characteristic peaks. Besides, preliminary pattern recognition had verified the chemometrics analytical results. Absolute peak area (APA) was used for relevant quantitative analysis, results showed the differences between two varieties and it was valuable for further quality control as selection of characteristic components.
Chromatography, High Pressure Liquid ; Citrus ; chemistry ; classification ; Discriminant Analysis ; Drugs, Chinese Herbal ; chemistry ; Fruit ; chemistry ; classification ; Mass Spectrometry ; Principal Component Analysis

OBJECTIVE: To establish the electricity plate digestion and inductively coupled plasma-mass spectrometry (ICP-MS) method for determination of 33 inorganic elements in human hair. METHODS: Lithium (6Li), Germanium (72Ge), Yttrium (89Y), Indium (115In), and Terbium (159Tb) were used as internal standards. The electric heating board digestion in a mixture of nitric acid and hydrogen peroxide was used as the pre-treatment of the hair. Thirty-three inorganic elements in human hair were analyzed by ICP-MS method. RESULTS: The detection limit of ICP-MS was 0.0001 microg/g(Th)-10.9 microg/g (Ca) and the limit of quantitation was 0.0005 microg/g (Th)-25 microg/g (Ca). The recovery rate of this method was 86%-113%. The RSD for the intra-day and inter-day were less than 9.2%. The method was not statistically different from microwave digestion method. CONCLUSION This method is highly efficient and accurate. It can be used for analysis of 33 inorganic elements in human hair.
Electricity ; Hair/chemistry* ; Humans ; Limit of Detection ; Mass Spectrometry/methods* ; Microwaves ; Reference Standards ; Sensitivity and Specificity ; Trace Elements/analysis*

OBJECTIVE: To determine the normal reference values of 33 elements, Ag, Al, As, Au, B, Ba, Be, Ca, Cd, Co, Cr, Cs, Cu, Fe, Ga, Hg, Li, Mg, Mn, Mo, Ni, Pb, Rb, Sb, Se, Sr, Th, Ti, Tl, U, V, Zn and Zr, in the blood and urine samples from the general population in Sanmen County of Zhejiang province, a typical coastal area of eastern China. METHODS: The 33 elements in 272 blood and 300 urine samples were determined by inductively coupled plasma-mass spectrometry (ICP-MS). The normality test of data was conducted using SPSS 17.0 Statistics. The data was compared with other reports. RESULTS: The normal reference values of the 33 elements in the blood and urine samples from the general population in Sanmen County were obtained, which of some elements were found to be similar with other reports, such as Co, Cu, Mn and Sr, while As, Cd, Hg and Pb were generally found to be higher than those previously reported. There was a wide variation between the reports from different countries in blood Ba. CONCLUSION The normal reference values of the 33 elements in the blood and urine samples from the general population in Sanmen County are established, and successfully applied to two poisoning cases.
Blood Chemical Analysis ; China ; Elements ; Environmental Monitoring ; Humans ; Urinalysis

ObjectiveTo develop school refusal reason inventory (SRRI)for children and adolescents in China and assess its reliability and validity.MethodsThe primary SSRI was made based on clinical interviews and literatures.Pretest was carried out in a small sample from a clinic.Then the final SSRI was developed after qualitative analysis and item analysis.SRRI,the Screen for Child Anxiety Related Emotional Disorders(SCARED) and Child Depression Inventory(CDI) were administered to school refusers from 7 schools in Shenyang.All the schools were selected from Shenyang City and its countryside by cluster sampling.Some of the students were retested after one month.Descriptive statistics and exploratory factor analysis were carried out to examine the reliability and validity of SRRI based on all the data.Results Item analysis indicated correlation coefficients between all the items and the total marks were higher than 0.3,and they were significant.All the critical ratios of the items were higher than 0.3.The 43 items were divided into six factors ( educational modality,factor of teachers,relationship with classmates,separated anxiety,study attitude and study environment) by exploratory factor analysis.The factor loading values were 0.372 ～0.848.The cronbach's α of each factor was 0.827,0.831,0.759,0.623,0.821 and 0.808.Retest reliability was 0.644 (P ＜ 0.01 ).Its correlation coefficient with SCARED was 0.452 and 0.548 with CDI.ConclusionAccording to Chinese cultural back ground,the SSRI corresponds with psychometric indexes.There are good reliability and validity.It is helpful to understand the reasons of school refusal behavior in children and adolescents.

To construct knockout vectors containing ampicillin resistant gene and partial sequence of hyaluronidase gene(Hyl)so that Hyl can be knock out by transforming the plasmid into Streptococcus zoopidemics mutans.First,partial sequence of Hyl(Hyl-1)was cloned into the vector of pMD19-T by using DNA of Streptococcus zoopidemics as template,and then a knockout vector pMD19T-SA was constructed,in which Hyl-1 gene was disrupted by inserting ampicillin resistant gene(Amp)from reverse PCR.As expected,the vector was proved to be consisted of Hyl-1-Amp-Hyl-1-pMD19-T.Thereafter,DNA fragment of Hyl-1-Amp-Hyl-1 was subcloned into pBR322 vector,the resulting construct was then checked by PCR and restriction analysis for the proper configuration of the knockout vector pBR322-SA.Both of the knockout vectors were used to transform Streptococcus zoopidemics and one recombinant was obtained in result.From results of PCR and Hyl activity assay,it was indicated that in the recombinant the Hyl gene was disrupted completely.

Background Our previous study demonstrated that curcumin can induce the apoptosis of retinal pigment epithelial (RPE) cells and herein inhibit the proliferation of RPE cells,and it is proved that the intravitreous injection of 0.1mg curcumin has less adverse effect to ocular tissue, inferring a good applicative prospect in clinic. Objective The goal of this experiment was to evaluate the effectiveness of curcumin on the prevention and treatment of experimental proliferative vitreoretinopathy (PVR). Methods PVR models were induced by injection of 0.1ml RPE cells (containing 2×106 cells) into vitreous cavity in 40 eyes of 20 healthy and mature New Zealand albino rabbits.0. 1ml curcumin(0. 1 mg) was then injected into lateral eye of each model rabbit immediately following the injection of RPE cells,and the equal volume of normal saline solution containing 0. 5‰ DMSO was injected into the fellow eye of each model rabbit as controls. On 1,3,7,14,21 and 28 days after injection, the changes of cornea, aqueous humor, lens, vitreous and fundus were examined and recorded by slit lamp biomicroscope, indirect ophthalmoscope,fundus color camera and B-type ultrasonograph to evaluate the inflammatory response. The incidence rate of retinal detachment was calculated and compared between curcumin group and control group. Results The inflammatory reaction in anterior chamber and misty opacity in vitreous were found from 1 day through 3 days after injection, but no obvious proliferative strap and retinal detachment in all of the experimental eyes. On the 7th day after injection, inflammatory reaction was extinct in the anterior chamber of rabbit eyes, and proliferative strap occurred in 14 eyes(75% ) in the control group but only 2 eyes (10% ) in curcumin group,showing significant difference between these two groups (P<0. 01). No retinal detachment was seen in both the two groups. On 14,21 and 28 days after injection, the incidence rate of retinal detachment was 55% ,80% ,95% respectively in control group and that of curcumin group was 10% ,15% ,15% respectively,presenting considerably differences between two groups (P<0. 01, P<0. 01 ,P<0. 01 ). Conclusion Injection of curcumin into vitreous cavity can effectively inhibit the occurrence and development of PVR in rabbit.