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 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

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 analyze the clinical characteristics and recent curative effect of mantle cell lymphoma (MCL) after conventional treatment.Methods Clinical data of 15 MCL patients admitted in the Affiliated Hospital of Academy of Military Medical Sciences between August 2004 and October 2013 were retrospectively analyzed.Results The median age of those patients was 59 and the male to female ratio was 1.5∶1.Fourteen(93%)cases were in Ann-Arbor stages Ⅲ -Ⅳ, 15 cases (100%)primarily with lymph node involvement,7 cases (47%)with bone marrow involvement,4 cases (27%)with gastrointestinal involvement,and 3 cases (20%)with orbit involvement.Less than 40% expression of Ki-67 was observed in 9 cases (60%),while 6 cases were with more than 40% (40%).One case was blastic variant.First-line therapy was CHOP-like regimens,which were combined with rituximab in 8 of the 15 cases.In this study,the median survival time was 12 months (3 -64),and the overal response rate was 80%after induction chemotherapy.The current survival of 7 /9 cases with less than 40% expression of Ki-67 was 8 -64 months,2 /6 cases with more than 40% expression of Ki-67 was 8 and 9 months,respectively.Conclusion MCL mostly occurs in older males.Extranodal invasion is common in MCL as an aggressive tumor.The efficacy of traditional chemotherapy is currently limited.Blastic variant or high expression of Ki-67 is an adverse prognostic indicator.

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.