利用炔基探针研究DNA结构动力学

外源振动探针的引入对生物分子特定区域的结构动力学研究具有重要意义。已有报道,较小的化学基团,如—N3和—CN,可作为跟踪探测DNA局部位点构象变化的振动探针。炔基,由于其对溶剂环境具有一定的敏感性,或许也可以成为分子局部结构的振动探针。而且,近期工作表明[1],在特定环境下,炔基的红外活性可被大幅度地增强,其跃迁偶极矩甚至有可能接近C≡O基团。在本工作中,我们将炔基修饰在DNA单体的碱基上,利用傅里叶变换红外光谱和飞秒二维红外光谱(2DIR),结合量化计算,研究了脱氧核糖核苷单体的结构动力学。

Nucleotide-binding Oligomerization Domain-1 Ligand Induces Inflammation and Attenuates Glucose Uptake in Human Adipocytes

Objective To investigate the effects of stimulant for nucleotide-binding oligomerization domain 1 （NOD1） on secretion of proinflammatory chemokine/cytokines and insulin-dependent glucose uptake in human differentiated adipocytes. Methods Adipose tissues were obtained from patients undergoing liposuction. Stromal vascular cells were extracted and differentiated into adipocytes. A specific ligand for NOD1, was administered to human adipocytes in culture. Nuclear factor-κB transcriptional activity and proinflammatory chemokine/cytokines production were determined by reporter plasmid assay and enzyme-linked immunosorbent assay, respectively. Insulin-stimulated glucose uptake was measured by 2-deoxy-D-[ 3 H] glucose uptake assay. Furthermore, chemokine/cytokine secretion and glucose uptake in adipocytes transfected with small interfering RNA （siRNA） targeting NOD1 upon stimulation of NOD1 ligand were analyzed. Results Nuclear factor-κB transcriptional activity and monocyte chemoattractant protein-1 （MCP-1）, interleukin （IL）-6, and IL-8 secretion in human adipocytes were markedly increased stimulated with NOD1 ligand （all P〈0.01）. Insulin-induced glucose uptake was decreased upon the activation of NOD1 （P〈0.05）. NOD1 gene silencing by siRNA reduced NOD1 ligand-induced MCP-1, IL-6, and IL-8 release and increased insulin-induced glucose uptake （all P〈0.05）. Conclusion NOD1 activation in adipocytes might be implicated in the onset of insulin resistance.

Genomic imbalances in esophageal carcinoma cell lines involve Wnt pathway genes

AIM： To identify molecular markers shared across South African esophageal squamous cell carcinoma （ESCC） cell lines using o/togenetics, fluorescence in situ hybridization （FISH） and single nucleotide polymorphism （SNP） array copy number analysis. METHODS： We used conventional cytogenetics, FISH, and multicolor FISH to characterize the chromosomal rearrangements of five ESCC cell lines established in South Africa. The whole genome copy number profile was established from 250K SNP arrays, and data was analyzed with the CNAT 4.0 and GISTIC software. tions involved the following chromosomal regions and genes： 11q13.3 （CCND1, FGF3, FGF4, FGF19, MYEOV）, 8q24.21（C-MYC, FAM84B）, 11q22.1-q22.3 （B[RC2, BIRC3）, 5p15.2 （CTNND2）, 3qll.2-q12.2 （MINA） and 18p11.32 （TYMS, YES1）. The significant deletions included 1p31.2-p31.1 （CTH, GADD45a, DIRAS3）, 2q22.1 （LRPIB）, 3p12.1-p14.2 （FHIT）, 4q22.1-q32.1 （CASP6, SMAD1）, 8p23.2-q11.1 （BNIP3L） and 18q21.1-q21.2 （SMAD4, DCC）. The 3p11.2 translocation breakpoint was shared across four cell lines, supporting a role for genes involved at this site, in particular, the EPHA3 gene which has previously been reported to be deleted in ESCC.CONCLUSION： The finding that a significant number of genes that were amplified （FGF3, FGF4, FGF19, CCND1 and C-MYC） or deleted （SFRP2 gene） are involved in the Wnt and fibroblast growth factor signaling pathways, suggests that these pathways may be activated in these cell lines.

Altofrequency SNPs of mitochondrial DNA in 26 Han Chinese

Objective: To explore the possible mitochondrial DNA (mtDNA) polymorphism in Han Chinese. Methods: The complete mitochondrial genome of 26 unrelated healthy Han Chinese were extracted and sequenced. Results:The mtDNA nucleotide sites (2 706, 7 028, 8 860, 11 719, and 15 326) were found totally different from the Revised Cambridge Reference Sequence (rCRS). These single nucleotide polymorphisms (SNPs) were 2 706 A→G, 7 028 C→T, 8 860 A→G, 11 719 G→A, 15 326 A→G. Conclusion: These findings provide new insights into the characteristics of Han Chinese mitochondrial genetic diversity.

Association of Interleukin-10 Gene Haplotype with Gastric Cancer in a Chinese Population

OBJECTIVE Interleukin-10 （IL-10） is a multifunctional cytokine with both immunosuppressive and antiangiogenic functions.Previous studies have reported that IL-10 levels are signifi cantly elevated in patients with gastric cancer （GC）. It has also been confirmed that interindividual variations in IL-10 production are genetically attributed to the polymorphisms of IL-10 gene.Therefore, this study was designed to investigate whether the polymorphisms of IL-10 gene were associated with susceptibility to GC in the Chinese population.METHODS The serum levels of IL-10 were measured by radioimmunoassay. The single nucleotide polymorphisms （SNPs） at positions -1082A/G, -819T/C and -592A/C in the IL-10 gene promoter were analyzed using polymerase chain reactionrestriction fragment length polymorphism （PCR-RFLP）.RESULTS 220 patients with gastric cancer and 180 healthy controls were included in this study. The serum levels of IL-10 were signifi cantly higher in GC patients than healthy controls （Z = -19.13, P 〈 0.001）. Single SNP analysis showed that the -1082G allele, -1082AG and -819CC genotypes significantly increased in patients with GC （P = 0.029, 0.021, 0.039 respectively）. In a logistic regression analysis adjusted for age and sex, the -1082AG genotype was associated with an odds ratio of 1.974 （95% CI,1.14-3.391; P = 0.014）, and the -819CC genotype with an odds ratio of 2.496 （95% CI, 1.222-5.102; P = 0.012） for GC. Furthermore,haplotype analysis revealed that at least five haplotypes （ATA,ACC, GCC, ACA and ATC） were existent in this population.Also that the GCC haplotype was associated with a signifi cantly increased risk of GC as compared with the ATA haplotype （OR =1.90; 95% CI, 1.11-3.27; P = 0.02）.CONCLUSION The results indicate that the gene haplotype of IL-10 may contribute to the susceptibility to GC in the Chinese population.

Variants of the Mitochondrial Displacement Loop in Patients with Myelodysplastic Syndromes

OBJECTIVE Some mtDNA mutations have been detected in patients with myelodysplastic syndromes(MDSs).As the non-coding region of mitochondria,the displacement loop(D-loop) region of mtDNA contains important elements for mtDNA replication and transcription.Variants of the D-loop region were found to be related to the cause of many diseases.The aim of our study was to investigate mutations and single nucleotide polymorphisms in the D-loop region of MDS patients.METHODS The mutations and SNPs in the hypervariable regions of the D-loop were detected by direct sequencing in MDS patients and normal controls.RESULTS Sixty-four SNPs were found in the D-loop region in MDS cases and control group.Among the SNPs,the 16,189 variant(T > C transition) was found to have an increased frequency in the MDS group(P = 0.044).However,no mutations were detected in neither group.CONCLUSION Our data provide evidence for a highly polymorphic D-loop region in patients with MDS,but do not support the presence of mutations in the mitochondrial D-loop region in MDS cases.The mtDNA T16,189C variant,which may be a functional variant,is associated with increased susceptibility to a MDS.

Variation of DRB1 Gene in Tibetan Sheep

It reveals that the MHC （major histocompatibility complex） gene product always involved in the control of immune response and disease resistance. Nowadays many studies have indicated the OLA （ovine lymphocyte antigen） DRB1 gene is associated with some sheep diseases. Tibetan sheep is one of the three major shag sheep breeds in China, and also have the largest number of China＇s sheep breeds. But till now no report has been seen on studying DRB1 gene in Tibetan sheep of China. To understand the evolution and provide the basis for sheep disease resistance, polymorphism in the exon2 ofDRB1 gene in Tibetan sheep was analyzed. The PCR-SSCP, cloning and sequencing were used to analyse DRB1 gene variation in 600 Tibetan sheep of China. And the genetic relationship and evolutionary significance of the alleles had also been analyzed. Total of 31 alleles were identified, in which 15 alleles had not been reported before. And there were 70 SNPs （single nucleotide polymorphisms） sites in 31 sheep DRB1 gene haplotypes, the proportion was 29.5% to the whole exort2 sequence. All of this indicated that DRB1 exon2 is highly polymorphic in Tibetan sheep. The variation identified here might have an impact on both the function and level of expression of the OLA-DRB1.

磷酰胺吗啉代寡核苷酸关键中间产物的合成方法探索与工艺优化

目的探索与优化磷酰胺吗啉代寡核苷酸(PMO)关键中间产物7'-羟基-N-三苯甲基吗啉代核苷单体合成工艺路线和方法,使其合成更加高效与便捷,为以PMO为结构基础的反义寡核苷酸类药物研究奠定基础。方法以N4-苯甲酰基胞苷,鸟苷与5-甲基尿苷作为起始原料,经过将核糖环结构改造为吗啉环并对关键化学基团保护等步骤合成7'-羟基-N-三苯甲基吗啉代核苷单体。结果分别得到N4-苯甲酰基-7'-羟基-N-三苯甲基吗啉代胞苷、N2-苯甲酰基-7'-羟基-N-三苯甲基吗啉代鸟苷和7'-羟基-N-三苯甲基吗啉代胸苷,并对三者合成的工艺方法进行了优化,对所合成的中间体和目标物进行了结构确证。结论优化后的7'-羟基-N-三苯甲基吗啉代核苷单体合成工艺高效、便捷且适合放大制备。

Association between NMDA receptor subunit 2b gene polymorphism and Alzheimer’s disease in Chinese Han population in Shanghai

Objective N-methyl-D-aspartate（NMDA）receptor has been indicated to be involved in the pathogenesis of Alzheimer’s disease（AD）.The NMDA receptor subunit 2b（NR2B）has attracted more attention due to its characteristic distribution and selective reduction in AD brain.The present study aimed to explore the association between NMDA gene polymorphism and AD.Methods A total of 63 AD patients and 68 normal controls in Shanghai city were employed in this study.Genotype of C2664T variant（rs1806201）in the exon13 of GRIN2B gene was determined by gene sequencing. Results Among AD patients,15（23.6%）subjects were identified as C/C genotype,and 35（55.6%）were identified as C/T genotype.The left 13（20.6%）subjects were identified as T/T genotype.In normal controls,15（22.1%）subjects were identified as C/C genotype,39（57.4%）as C/T genotype and 14（20.6%）as T/T genotype.The distribution frequency of neither GRIN2B C2664T genotype（P=0.895）nor allele（P=0.790）was significantly different between AD patients and normal controls,even when the subjects were stratified by gender and age of disease onset in AD patients.Conclusion The results suggest that there is no relation between GRIN2B C2664T polymorphism and AD in Chinese Han population of Shanghai City.

Polymorphisms of the IGF1 gene and their association with growth traits, serum concentration and expression rate of IGF1 and IGF1R in buffalo

The insulin-like growth factor 1（IGF1） gene is a member of the group of somatotropin axis genes that play a significant role in cell proliferation and growth of muscles. Here, we searched for polymorphisms in buffalo IGF1 and found two novel single nucleotide polymorphisms（SNPs）, G64 A and G280A, in the noncoding sequences of exon 1 and exon 4, respectively. Statistical analysis of different genotypes showed that the individuals with GG genotypes had significantly（P〈0.05） higher body weight（BW） and average daily gain（ADG） than those with other genotypes at ages of 3–6 months in G64A SNP and 6–9 months in G280A SNP. The combined genotypes of these two SNPs produced three haplotypes, GG/GG, AG/AG, and AA/AA, which were significantly associated（P〈0.0001） with BW and ADG at an age from 3 to 12 months. Buffaloes with the homozygous GG/GG haplotype showed higher growth performance than other buffaloes. The two SNPs were correlated with m RNA levels of IGF1 and IGF1 receptor（IGF1 R） in semitendinosus muscle as well as with the serum concentration level of IGF1. Also, buffaloes with GG/GG haplotype showed higher m RNA and serum concentration levels. The data revealed that these two SNPs could be valuable genetic markers for selection of Egyptian buffaloes for better performance in the population.

De novo identification and quantification of single amino-acid variants in human brain

The detection of single amino-acid variants （SAVs） usually depends on single-nucleotide polymorphisms （SNPs） database. Here, we describe a novel method that discovers SAVs at proteome level independent of SNPs data. Using mass spectrometry-based de novo sequencing algorithm, peptide-candidates are identified and compared with theoretical protein database to generate SAVs under pairing strategy, which is followed by database re-searching to control false discovery rate. in human brain tissues, we can confidently identify known and novel protein variants with diverse origins. Combined with DNA/RNA sequencing, we verify SAVs derived from DNA mutations, RNA alternative splicing, and unknown post-transcriptional mechanisms. Furthermore, quantitative analysis in human brain tissues reveals several tissue-specific differential expressions of SAVs. This approach provides a novel access to high-throughput detection of protein variants, which may offer the potential for clinical biomarker discovery and mechanistic research.

Haplotype of platelet receptor P2RY12 gene is associated with residual clopidogrel on-treatment platelet reactivity

Objective： To investigate a possible association between common variations of the P2RY12 and the residual clopidogrel on-treatment platelet reactivity after adjusting for the influence of CYP2C19 tested by thromboe- lastography （TEG）. Methods： One hundred and eighty patients with acute coronary syndrome （ACS） treated with clopJdogrel and aspJdn were included and platelet function was assessed by TEG. Five selected P2RY12 single nu- cleotide polymorphisms （SNPs; rs6798347, rs6787801, rs6801273, rs6785930, and rs2046934）, which cover the common variations in the P2RY12 gene and its regulatory regions, and three CYP2C19 SNPs （ 2, 3, 17） were geno- typed and possible haplotypes were analyzed. Results： The high on-treatment platelet reactivity （HTPR） prevalence defined by a platelet inhibition rate 〈30% by TEG in adenosine diphosphate （ADP）-channel was 69 （38.33%）. Six common haplotypes were inferred from four of the selected P2RY12 SNPs （denoted H0 to H5） according to the linkage disequilibrium R square （except for rs2046934）. Haplotype H1 showed a significantly lower incidence of HTPR than the reference haplotype （H0） in the total study population while haplotypes H1 and H2 showed significantly lower incidences of HTPR than H0 in the nonsmoker subgroup after adjusting for CYP2Clg effects and demographic characteristics. rs2046934 （T744C） did not show any significant association with HTPR. Conclusions： The combination of common P2RY12 variations including regulatory regions rather than rs2046934 （T744C） that related to pharmacodynamics of clopidogrel in patients with ACS was independently associated with residual on-clopidogrel platelet reactivity. This is apart from the established association of the CYP2C19. This association seemed more important in the subgroup defined by smoking.

Identification of a combination of SNPs associated with Graves' disease using swarm intelligence

Graves' disease,the production of thyroid-stimulating hormone receptor-stimulating antibodies leading to hyperthyroidism,is one of the most common forms of human autoimmune disease.It is widely agreed that complex diseases are not controlled simply by an individual gene or DNA variation but by their combination.Single nucleotide polymorphisms(SNPs),which are the most common form of DNA variation,have great potential as a medical diagnostic tool.In this paper,the P-value is used as a SNP pre-selection criterion,and a wrapper algorithm with binary particle swarm optimization is used to find the rule for discriminating between affected and control subjects.We analyzed the association between combinations of SNPs and Graves' disease by investigating 108 SNPs in 384 cases and 652 controls.We evaluated our method by differentiating between cases and controls in a five-fold cross validation test,and it achieved a 72.9% prediction accuracy with a combination of 17 SNPs.The experimental results showed that SNPs,even those with a high P-value,have a greater effect on Graves' disease when acting in a combination.

Bivariate genome-wide association study suggests that the DARC gene influences lean body mass and age at menarche

Lean body mass （LBM） and age at menarche （AAM） are two important complex traits for human health. The aim of this study was to identify pleiotropic genes for both traits using a powerful bivariate genome-wide association study （GWAS）. Two stud- ies, a discovery study and a replication study, were performed. In the discovery study, 909622 single nucleotide polymor- phisms （SNPs） were genotyped in 801 unrelated female Han Chinese subjects using the Affymetrix human genome-wide SNP array 6.0 platform. Then, a bivariate GWAS was performed to identify the SNPs that may be important for LBM and AAM. In the replication study, significant findings from the discovery study were validated in 1692 unrelated Caucasian female subjects One SNP rs3027009 that was bivafiately associated with left arm lean mass and AAM in the discovery samples （P=7.26x10-6） and in the replication samples （P=0.005） was identified. The SNP is located at the upstream of DARC （Duffy antigen receptor for chemokines） gene, suggesting that DARC may play an important role in regulating the metabolisms of both LBM and AAM.

SCANNING AND ANALYSIS OF MISMATCH DISTRIBUTION ON HUMAN GENOME

The mismatch distribution is a good descriptive summary statistic that describes the phenomena of population genetics. This article scanned mismatch distribution on human genome with single nucleotide polymorphism （SNP） data from the International HapMap Project. It is found that the abnormal mismatch distribution could imply some special segments on some chromosomes. One of the segments, on chromosome 8, was proved as an inversion. Other special segments may also imply some special structure on chromo- somes, such as duplication. The conjectures of other segments still need further research.