Impact of cognition-related single nucleotide polymorphisms on brain imaging phenotype in Parkinson’s disease

Multiple single nucleotide polymorphisms may contribute to cognitive decline in Parkinson’s disease. However, the mechanism by which these single nucleotide polymorphisms modify brain imaging phenotype remains unclear. The aim of this study was to investigate the potential effects of multiple single nucleotide polymorphisms on brain imaging phenotype in Parkinson’s disease. Forty-eight Parkinson’s disease patients and 39 matched healthy controls underwent genotyping and 7 T magnetic resonance imaging. A cognitive-weighted polygenic risk score model was designed, in which the effect sizes were determined individually for 36 single nucleotide polymorphisms. The correlations between polygenic risk score, neuroimaging features, and clinical data were analyzed. Furthermore, individual single nucleotide polymorphism analysis was performed to explore the main effects of genotypes and their interactive effects with Parkinson’s disease diagnosis. We found that, in Parkinson’s disease, the polygenic risk score was correlated with the neural activity of the hippocampus, parahippocampus, and fusiform gyrus, and with hippocampal-prefrontal and fusiform-temporal connectivity, as well as with gray matter alterations in the orbitofrontal cortex. In addition, we found that single nucleotide polymorphisms in α-synuclein(SNCA) were associated with white matter microstructural changes in the superior corona radiata, corpus callosum, and external capsule. A single nucleotide polymorphism in catechol-O-methyltransferase was associated with the neural activities of the lingual, fusiform, and occipital gyri, which are involved in visual cognitive dysfunction. Furthermore, DRD3 was associated with frontal and temporal lobe function and structure. In conclusion, imaging genetics is useful for providing a better understanding of the genetic pathways involved in the pathophysiologic processes underlying Parkinson’s disease. This study provides evidence of an association between genetic factors, cognitive functions, and multi-modality neuroimaging biomarkers in Parkinson’s disease.

A single nucleotide polymorphism in XRCC4 gene is associated with reduced colorectal cancer susceptibility in female

Objective： To investigate the association of XRCC4 polymorphic variants at G-1394T （rs6869366） with colorectal cancer susceptibility. Methods： In this hospital-based case-control study, the association of XRCC4 polymorphism with colorectal cancer risk in Chinese population was investigated. In total, 171 patients with colorectal cancer and 171 healthy individuals matched for age and gender were selected. The genomic DNAs of the patients and controls were extracted from peripheral blood and the 300 bp target DNA was amplified with Polymerase Chain Reaction. The products were then digested with restriction endonuclease HinclI, followed by agarose electrophoresis to identify the genotype. Results： We found a significant difference in the frequency of the XRCC4 G-1394T genotype between the colorectal cancer and control groups in female （1/127 vs 8/122, P〈0.05）. Those with G/T at XRCC4 G-1394T showed a decreased risk of colorectal cancer susceptibility compared with those with T/T （OR 0.113, 95%CI 0.014-0.932）. However, in overall population or in male, there was no significant difference of the distribution between the colorectal cancer and control groups. Conclusion： Our findings with decreased risk of colorectal cancer susceptibility suggested that the G allele of XRCC4 G-1394T were associated in female.

Development of organelle single nucleotide polymorphism (SNP) markers and their application for the identification of cytoplasmic inheritance patterns in Pyropia yezoensis (Bangiales,Rhodophyta)

The genus Pyropia contains several important cultivated species.Genetic research in nori species has mainly focused on the cell nucleus,with few studies on organelles(chloroplast and mitochondria).Due to the high copy numbers of organelles in cells,which influence the development and traits of algae,it is necessary to study their genetic mechanism.In this study,the marine red alga Pyropia yezoensis,an important economic macroalga,was selected as the study object.To investigate organelle(chloroplast and mitochondria)inheritance in P.yezoensis,the wild type RZ(maternal strain)was crossed with the red mutant HT(paternal strain)and 30 color-sectors from 11 F1 gametophytic blades were examined.The complete chloroplast and mitochondrial genomes of the red mutant(HT)were assembled for the first time.One reliable and stable single nucleotide polymorphism(SNP)loci filtrated by bioinformatics analysis was used as a molecular marker for chloroplast and mitochondrial DNA,respectively,in subsequent experiments.PCR amplification and sequence analysis showed that the haplotypes of color-sectors detected were consistent with those of the maternal parent,confirming that both chloroplast and mitochondrial genomes were inherited maternally in P.yezoensis.The inheritance pattern of organelles in P.yezoensis can be used to guide the hybridization and breeding of nori.Additionally,the organelle SNP markers developed in this study can be applied in subsequent genetic research.

Relation of cytochrome P450 2C19 gene 681G>A single nucleotide polynmrphism to clopidogrel resistance after PCI in Chinese

Objectives Clopidogrel is a prodrug that has to be converted to an active metabolite by hepatic cytochrome P450(CYP) isoenzymes to inhibit platelet aggregation.Individualvariability of platelet inhibition by clopidogrel suggests a possibility for genetic factors having a significant influence on clopidogrel responsiveness.In this study,we sought to determine the association between the single nucleotide polymorphism of CYP 2C19 681G】A and the occurrence of clopidogrel resistance(CR) in Chinese.Methods The study enrolled 614 hospitalized patients who underwentsuccessful percutaneouscoronary intervention with drug-eluting stents were received the treatmentwith dual antiplatelet regimen(aspirin plus clopidogrel).All patients received loading doses of 600 mg clopidogrel and 300 mg aspirin.20μmol/L ADP-induced platelet aggregation ratio(PAR ) was assessed 24 h after clopi- dogrel administration.The maximum residual PAR≥70%was defined as CR.Genomic DNA was extracted from whole blood samples according to standard protocols,the single nucleotide polymorphism of the CYP2C19 681G】A was genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in all the patients.Results CR was found in 126 patients(20.5%).There was CYP2C19 681G】A polymorphism in the study population.The frequencies of the three kinds of genotypes(GG,GA,A A) in CR group and non-CR (NCR)group were 32.5%,47.6%,19.8%and 48.0%, 45.0%,7.0%,respectively.The frequency of AA genotype was significantly higher in NCR group than that in CR group (OR =3.03,95%CI:1.889～5.784,P=0.003).The A allele carriers were more likely to develop clopidogrel resistance compared with that of G allele carriers(OR=1.85,95%CI: 1.392～2.459,P=0.002).Conclusions CYP2C19 681G/A polymorphism is associated with the risk of CR,and the A allele carriers may be a possible genetic susceptibility factor for patients with CR.

Identification of a Regulatory Single Nucleotide Polymorphism in the Adiponectin (APM1) Gene Associated with Type 2 Diabetes in Han Nationality

Objective To identify the genetic defects of the the adiponectin （APM1） gene that contribute to the development of type 2 diabetes （T2DM） and determine the functional single nucleotide polymorphisms （SNPs） in the APMI gene associated with T2DM in Han nationality. Methods The APMI gene 5＇-UTR was screened by direct sequencing to identify common polymorphisms. Identified SNPs were genotyped in 585 nondiabetic controls, 278 subjects with impaired glucose intolerance （IGT） and 212 patients with T2DM. The functions of SNPs in the regulatory region were assessed by reporter gene assay. Possible association between SNPs and plasma APMI levels or metabolic parameters was statistically asses,sed. Results Three SNPs were identified in the APMI gene 5＇-UTR. A case-control study revealed that SNP -11377 G/C had significant differences in allele frequencies between T2DM patients and nondiabetic controls （G 0.314/C 0.686 vs. G 0.265/C 0.735, P=0.03）. Haplotype analysis of three SNPs in the APM1 gene showed that no significant association of haplotypes with T2DM. IGT was detected in the present study. Reporter gene assay showed that SNP did not influence the transcription efficiency in the 3T3-LI cell line. Conclusion SNP - 11377 G/C in the proximal promoter region of the APM 1 gene contributes to the development of T2DM in Han nationality but may not be a functional SNP in the APM1 gene.

Gene-gene,gene-environment,gene-nutrient interactions and single nucleotide polymorphisms of inflammatory cytokines

Inflammation plays a significant role in the etiology of type 2 diabetes mellitus(T2DM).The rise in the pro-inflammatory cytokines is the essential step in glucotoxicity and lipotoxicity induced mitochondrial injury,oxidative stress and beta cell apoptosis in T2 DM.Among the recognized markers are interleukin(IL)-6,IL-1,IL-10,IL-18,tissue necrosis factor-alpha(TNF-α),C-reactive protein,resistin,adiponectin,tissue plasminogen activator,fibrinogen and heptoglobins.Diabetes mellitus has firm genetic and very strong environmental influence; exhibiting a polygenic mode of inheritance.Many single nucleotide polymorphisms(SNPs) in various genes including those of pro and antiinflammatory cytokines have been reported as a risk for T2 DM.Not all the SNPs have been confirmed by unifying results in different studies and wide variations have been reported in various ethnic groups.The inter-ethnic variations can be explained by the fact that gene expression may be regulated by gene-gene,gene-environment and gene-nutrient interactions.This review highlights the impact of these interactions on determining the role of single nucleotide polymorphism of IL-6,TNF-α,resistin and adiponectin in pathogenesis of T2 DM.

Association of three single nucleotide polymorphisms of ESR1 with breast cancer susceptibility:a meta-analysis

Expression of estrogen receptors is correlated with breast cancer risk,but inconsistent results have been reported.To clarify potential estrogen receptor（ESR）-related breast cancer risk,we analyzed genetic variants of ESR1 in association with breast cancer susceptibility.We performed a meta-analysis to investigate the association between rs2234693,rs1801132,and rs2046210（single nucleotide polymorphisms of ESR1）,and breast cancer risk.Our analysis included 44 case-control studies.For rs2234693,the CC genotype had a higher risk of breast cancer compared to the TT or CT genotype.For rs2046210,the AA,GA,or GA ＋ GG genotype had a much higher risk compared to the GG genotype.No significant association was found for the rs 1801132 polymorphism with breast cancer risk.This meta-analysis demonstrates association between the rs2234693 and rs2046210 polymorphisms of ESR1 and breast cancer risk.The correlation strength between rs2234693 and breast cancer susceptibility differs in subgroup assessment by ethnicity.

Association between low molecular polypeptide 7 single nucleotide polymorphism and response to therapy in hepatitis C virus infection

AIM: To investigate the relationship between low molecular polypeptide-7 (LMP-7) gene polymorphism and response to interferon (IFN) therapy in chronic hepatitis C virus (HCV) patients. METHODS: LMP-7 polymorphism at codon 49 with nucleotide substitution from A to C was amplified in 104 chronic HCV patients of genotype 4. The amplicons were digested with restriction endonuclease Bsm I and the produced restriction fragment length polymorphism was analyzed. Patients received IFN + regional blood volume therapy for 48 wk and the frequency of thissingle nucleotide polymorphism (SNP) was statistically correlated with treatment response. The exclusion criteria for these patients were stated by the national health program for treating viral hepatitis. Main exclusion criteria included co-infection with hepatitis B virus or schistosomiasis, thyroid dysfunction, uncontrolled diabetes mellitus, history of long term drug or alcohol intake and autoimmune hepatitis. Multivariate analyses were done to correlate LMP-7 SNP plus several factors such as age, gender, weight, serum alpha-fetoprotein (AFP) and alanine aminotransferase levels, liver activity, fibrosis score and viral load with response to therapy. RESULTS: The data presented in this study clearly demonstrated statistically significant differences between sustained virological response (SVR) (defined as the absence of HCV RNA levels in the patient's sera at least 6 mo after discontinuation of treatment) and non-response (NR) (where HCV RNA levels in the patient's sera never become undetectable for 6 mo during or after treatment). Variables were described as odds ratio with 95%CI. The data were considered significant if P values were ≤ 0.05; highly significant if P < 0.01 and very highly significant if P < 0.001. Current data showed that 91.7% of patients carrying LMP-7 C/C allele were associated with SVR, while the other two genotypes C/A and A/A were associated with NR patients, 83.3% and 64.3% respectively, showing that genotype CC was strongly associated with response to interferon (95%CI: 12.0719-134.6572, P = 0.0001). The majority of parameters recorded in SVR and NR patients included higher values of mean age (P = 0.004), alanine aminotransferase (P = 0.001), AFP (P = 0.001), body weight (P = 0.025), viral load (P = 0.025), higher fibrosis and histological activity index indices among NR vs SVR patients. Also, the multivariate statistical analysis of the different factors of fibro-sis score, liver activity grade, genotypes and alleles of LMP-7 gene polymorphism in responders and NRs of HCV patients in this study showed that HCV patients with A allele had a very highly significant association with the NRs, high fibrosis and higher liver activity, while the C allele had a very highly significant association with the responders, low fibrosis and lower liver activity (95%CI: 3.5800-13.2519, P = 0.0001).CONCLUSION: LMP-7 SNP is a candidate gene that should be considered when designing a mathematical model for predicting response to therapy and disease progression in HCV patients.

Role of matrix metalloproteinase,tissue inhibitor of metalloproteinase and tumor necrosis factor-α single nucleotide gene polymorphisms in inflammatory bowel disease

AIM:To study the (functional) relevance of single nucleotide polymorphisms (SNPs) in genes encoding matrix metalloproteinases (MMP)-1,-2,-3,-9,tissue inhibitors of metalloproteinases (TIMP)-1,-2 and tumor necrosis factor (TNF)-α in the etiopathogenesis of inflammatory bowel diseases (IBD),that may enhance susceptibility and/or disease severity. METHODS:Genomic DNA from 134 Crohn's disease (CD),111 ulcerative colitis (UC) patients and 248 control subjects was isolated from resected intestinal tissue or blood. Allelic composition at SNP loci was determined by PCR-RFLP or tetra primer ARMS PCR. RESULTS:The TIMP-1 genotype TT in women and T in men at SNP +372 T/C was found to increase CD susceptibility (39% vs 23.8%,P=0.018 and 67.9% vs 51.6%,P=0.055,respectively),while women with this genotype were less prone to development of fistulae during follow-up (41.4% vs 68.3%,P=0.025). Male IBD or CD patients carrying the TIMP-1 +372 T-allele expressed lower levels of TIMP-1 in surgically resected macroscopically inflamed tissue (0.065 < P < 0.01). The 5T5T genotype at MMP-3 SNP -1613 5T/6T increased the chance of stenotic complications in CD during follow-up (91.2% vs 71.8%,P = 0.022) but seemed to protect against colonic involvement of this disease at first endoscopic/radiologic examination (35.3% vs 59.5%,P=0.017). CONCLUSION:Allelic composition at the examinedSNPs in genes coding for TIMP-1 and MMP-3 affect CD susceptibility and/or phenotype,i.e.,fistulizing disease,stricture pathogenesis and first disease localisation. These findings reinforce the important role of these proteins in IBD.

Correlation between single nucleotide polymorphism of rs3811047 in IL-1 F7 gene and rheumatoid arthritis susceptibility among Han population in central plains of China

Objective:To discuss the association between single nucleotide polymorphism(SNP) of rs3811047 in IL-1 F7 gene and rheumatoid arthritis(RA) susceptibility among the Han population in central plains of China.Methods:A total of 276 RA patients admitted to our hospital from December 2009 to December 2011 together with 276 healthy physical examinees in the same period were chosen as the subjects.The typing for rs3811047 SNP in IL-1 F7 gene was carried out by using ligase detection reaction and polymerase chain reaction technique.And the frequency of each allele and genotypes distribution was calculated so as to evaluate the association between genotype distribution and RA susceptibility.Results:The frequency of A allele of rs3811047 in IL-1 F7 gene in RA group and control group was 16.27%and 17.68%,respectively,and that of G allele in two groups was 83.73%and 82.32%,respectively.The difference between two groups wasn’t statistical significant(P 】0.05).The frequency of genotype AA,AG and GG in RA group was 2.19%,27.84%and 69.97%,respectively,while that in control group was 2.94%,29.78%and 67.28%, respectively.The difference of distribution of three genotypes was not statistically significant (P 】0.05).RA patients with A allele were better than those without A allele in joint swelling index, rest pain,HAQ scoring and blood sedimentation.There was significant difference between two groups in above indexes(P【0.05/P【0.01).Conclusions:No significant correlation between RA susceptibility among the Han population in central plains of China and rs3811047 SNP inIL-1 F7 gene is observed.However,A allele of rs3811047 has certain influence on the condition of RA patients.

Investigation of HER-2 codon 655 single nucleotide polymorphism frequency and c-ErbB-2 protein expression alterations in gastric cancer patients

AIM： To investigate both whether the risk of gastric cancer is associated with the Ile/Val single nucleotide polymorphism （SNP） of human epidermal growth factor receptor-2 （HER-2） transmembrane domain-coding region at codon 655 and the suggested existence of HER-2 expression in gastric cancer cases in a Turkish patient group. METHODS： Polymerase chain reaction （PCR） followed by restriction fragment length polymorphism （RFLP） strategy was used to analyze the presence of HER-2 SNP at codon 655. c-erbB-2 expression pattern was analyzed by immunohistochemistry. The results were compared between gastric carcinoma group and chronic gastritis group, as well as between clinicopathological parameters and carcinoma. RESULTS： Results showed that Ile/Val genotype accounted for 20% within the Turkish gastric carcinoma group, and none in chronic gastritis group, and this genotyping was associated with stage Ⅳ gastric cancers （P = 0.04）. Positive membranous HER-2 immunoreactivity, on the other hand, accounted for 24% within the Turkish gastric carcinoma group and none from chronic gastritis cases; further, it was correlated with intestinal type carcinomas （P = 0.007）, and stage Ⅲ-Ⅳ carcinomas （P = 0.004）. CONCLUSION： These observations imply that the tested HER-2 SNP may participate in the development and progression of gastric cancer. Thus, after confirming these results with large sample groups, HER-2 codon 655 SNP and/or c-erbB-2 overexpression may also be used as a poor prognostic indicator for gastric carcinomas.

Tagging single nucleotide polymorphisms in the PPAR-γ and RXR-α gene and type 2 diabetes risk:a case-control study of a Chinese Han population

Peroxisome proliferator-activated receptor （PPAR-γ）,which is mainly involved in adipocyte differentiation, has been suggested to play an important role in the pathogenesis of insulin resistance and atherosclerosis. We investigated the frequencies of two common tagging polymorphisms of the PPAR-γ gene and two of PPAR-α with minor allele frequency （MAF）≥ 0.05 in the Chinese Han population and analyzed the correlation between the different genotypes and the risk of type 2 diabetes mellitus （T2DM）. TaqMan assay was performed to test the genotypes in T2DM patients （n = 1,105） and normal controls （n = 1,107）. Serum adiponectin concentration was measured by ELISA kit. The variant genotypes rs17817276GG, rs3856806CT and rs3856806CT/TT of PPAR-γ were associated with T2DM, P = 0.023,0.037 and 0.018, respectively. Furthermore, the prevalence of haplotype GT in PPAR-γ was less frequent in the case subjects （0.3%） than in the controls （1.9%） [P 0.001,OR（95%CI）=0.13 （0.06-0.31）]. Patients with genotype TT of rs3856806 had a higher serum level of adiponectin than those with the genotype CC and CT （P = 0.031 and 0.038, respectively）. There was no statistically significant difference between patients and controls in genotype distribution of rs6537944 and rs1045570 of the RXR-α gene. The present study suggests that the variant genotypes in the PPAR-γ gene could decrease the risk for the development of T2DM in the Chinese Han population.

Association of single nucleotide polymorphism of vitamin D receptor gene start codon and the suscepbility to prostate cancer in Han nationality in Hubei, China

Aim: To investigate the single nucleotide polymorphism of vitamin D receptor (VDR) gene start codon in the Han nationality in Hubei and its relationship to the susceptibility to prostate cancel (PCa). Methods: The VDR genotypes were determined by poly-merase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 80 patients with PCa and 96 normal male controls from the Han nationality in Hubei, using endonuclease Fok. Direct sequencing was done in part of the PCR products. Results: The frequency distribution of Fok I alleles in this cohort all followed the Hardy-Weinberg equilibrium. The distribution of genotypes and alleles had no significant difference between PCa patients and the controls (P>0.05). Conclusion: There was no significant relationship between Fok I polymorphism of VDR gene start codon and PCa in the Han nationality in Hubei.

Clinical significance of NOD2/CARD15 and Toll-like receptor 4 gene single nucleotide polymorphisms in inflammatory bowel disease

AIM： To evaluate the role of genetic factors in the pathogenesis of Crohn＇s disease （CD） and ulcerative colitis （UC）, we investigated the single nucleotide polymorphisms （SNPs） of NOD2/CARD15 （R702W, Gg08R and L1007finsC）, and Toll-like receptor 4 （TLR4） genes （D299G and T399I） in a selected inflammatory bowel disease （IBD） population coming from Southern Italy. METHODS： Allele and genotype frequencies of NOD2/ CARD15 （R702W, Gg08R and L1007finsC） and TLR4 （D299G and T399I） SNPs were examined in 133 CD patients, in 45 UC patients, and in 103 healthy controls. A genotype-phenotype correlation was performed. RESULTS： NOD2/CARD15 R702W mutation was significantly more frequent in CD （9.8%） than in controls （2.4%, P = 0.001） and in UC （2.3%, P = 0.03）. No significant difference was found between UC patients and control group （P 〉 0.05）. In CD and UC patients, no significant association with G908R variant was found. L1007finsC SNP showed an association with CD （9.8%） compared with controls （2.9%, P = 0.002） and UC patients （2.3%, P = 0.01）. Moreover, in CD patients, G908R and L1007finsC mutations were significantly associated with different phenotypes compared to CD wild-type patients. No association of IBD with the TLR4 SNPs was found in either cohort （allele frequencies： D299G-controls 3.9%, CD 3.7%, UC 3.4%, P 〉 0.05; T399I-controls 2.9%, CD 3.0%, UC 3.4%, P 〉 0.05）. CONCLUSION： These findings confirm that, in our IBD patients selected from Southern Italy, the NOD2/ CARD15, but not TLR4 SNPs, are associated with increased risk of CD.

Association of single nucleotide polymorphisms of brain-derived neurotrophic factor gene and multidrug resistance 1 gene to refractory epilepsy in Chinese Han children

BACKGROUND： There are two hypotheses for the underlying cause of refractory epilepsy： ＂target＂ and ＂transport＂. Studies have shown that brain-derived neurotrophic factor （BDNF） is over-expressed in refractory epilepsy. Multidrug resistance 1 （MDR1） gene encodes for P-glycoprotein, the primary ATP-binding cassette transporter in the human body. Some single nucleotide polymorphisms of the MDR1 gene have been associated with refractory epilepsy. OBJECTIVE： To investigate the association between BDNF gene C270T polymorphism and MDR1 T-129C polymorphism with refractory epilepsy in Chinese Han children through the use of polymerase chain reaction （PCR）-restriction fragment length polymorphism analysis. DESIGN, TIME AND SETTING： A case-control, genetic association study was performed at the Central Laboratory, Third Xiangya Hospital of Central South University from June 2005 to November 2007. PARTICIPANTS： A total of 84 cases of unrelated children with epilepsy, including 41 cases of refractory epilepsy and 43 cases of drug-responsive epilepsy, were enrolled. An additional 30 healthy, Chinese Han children, whose ages and gender matched the refractory epilepsy patients, were selected as normal controls. METHODS： Venous blood was collected and genomic DNA was extracted from the blood specimens. C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene were genotyped using PCR-restriction fragment length polymorphism analysis. Association analysis using the Ftest and Chi-square test was statistically performed between C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene and refractory epilepsy. MAIN OUTCOME MEASURES： The distribution of genotypes and allele frequencies of C270T polymorphism in BDNF gene and T-129C polymorphism in MDR1 gene. RESULTS： The distribution of CC, CT, and TT genotypes, as well as C and T allele frequencies, in the BDNF gene was not significantly different between the refractory epilepsy group, drug-responsive epilepsy group, or the normal control group （P 〉 0.05）. The distribution of TT genotype and T allele frequencies of the MDR1 gene was significantly different in the refractory epilepsy group compared with the drug-responsive epilepsy and normal control groups （P 〈 0.05）. Comparison of haplotype combinations demonstrated that there were no significant differences in combinations of TT＋CC, -FI-＋CT, TC＋CC, and TC＋CT among the three groups （P 〉 0.05）. CONCLUSION： C270T polymorphism of the BDNF gene was not associated with refractory epilepsy in Chinese Han children, but T-129C polymorphism in the MDR1 gene was associated with refractory epilepsy in Chinese Han children. The TT genotype and T allele frequencies could serve as susceptibility loci for refractory epilepsy. Interactions between C270T in BDNF gene and T-129C in MDR1 gene were not observed in refractory epilepsy in Chinese Han children.

Isolation,expression and single nucleotide polymorphisms(SNPs) analysis of LACCASE gene(LkLAC8) from Japanese larch(Larix kaempferi)

Nucleotide diversity (pi) and linkage disequilibrium (LD) analysis based on SNP marker could provide a sound basis for choosing an association analysis method. Japanese larch (Larix kaempferi) is an important timber coniferous tree species for pulping and papermaking, but its high lignin content has significantly restricted it application potential. In this study, the LACCASE gene, that plays an important regulatory role for lignin biosynthesis, was selected as research target. The full-length cDNA and genomic sequences of the encoding LkLAC8 gene were isolated from the LACCASE expressed sequence tags of the Japanese larch transcriptome database using the rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR). The cDNA was determined to be 1940 bp, with an open reading frame (ORF, 1734 bp) that encoded a protein of 577 AA. This protein contains four highly specific Cu2+ binding sites and 11 glycosylation sites, thus belonging to the LACCASE family. The deduced protein sequence shared an 89% identity with the PtaLAC from Pinus taeda. A real-time PCR analysis showed that the LkLAC8 transcript was expressed predominantly in mature xylem, with moderate levels in the immature xylem, cambium and mature leaves, the lowest in the roots. Lastly, the genomic sequences of LkLAC8 in 40 individuals from six naturally distributed populations of Japanese larch were amplified, and a total of 201 SNPs (103 and 98 mutation types of transition and transversion, respectively) were detected; the frequency of the SNPs was 1/19 bp. Nucleotide diversity among the six populations ranged from 0.0034 to 0.0053, which suggested that there were no significant differences among the populations. The LD analysis showed that the LD level decayed rapidly within the increasing length of the LkLAC8 gene. These results implied that LD mapping and association analysis based on candidate gene may be feasible for the marker-assisted breeding of new germplasms with low lignin in Japanese larch.

Relationship between Single Nucleotide Polymorphisms in -174G/C and -634C/G Promoter Region of Interleukin-6 and Prostate Cancer

The association between the single nucleotide polymorphisms （SNPs） in -174G/C and -634C/G of interleukin-6 （IL-6） promoter region and prostate cancer was examined in the population of Han people in Hubei region. TaqMan PCR was employed for the gene-typing of -174G/C and -634C/G in promoter region of IL-6 gene to compare the prostate cancer patients and normal controls in terms of genotype frequency, allele frequency and risk of prostate cancer. Enzyme-linked immu- nosorbent assay （ELISA） was used for the detection of IL-6 concentration in peripheral blood of the patients with prostate cancer and the relationship between the IL-6 level and the genotype was studied. Our results showed that in all the subjects, the genotype of genetic locus -174G/C was found to be GG and no CG and CC were observed. There was a significant difference in gene frequency of GG, CG and CC of -634C/G and allele frequency of G and C between prostate cancer patients and normal controls （P〈0.05） and the gene frequency of GG＋CG increased with the clinical stages and pathological grades of prostate cancer. The IL-6 level in GG＋CG group was significantly higher than that in CC group. It was concluded that no SNP in -174G/C IL-6 promoter region was found in the population of Han people in Hubei region. The SNP in -634C/G was, to some extent, associated with the development and progression of prostate cancer. The population with GG＋CG genetype has higher risk for prostate cancer.

A Novel Real-time Fluorescence Mutant-allele-specific Amplification Method for Rapid Single Nucleotide Polymorphism Analysis

Current methods for single nucleotide polymorphism （SNP） analysis are timeconsuming and complicated. We aimed at development of one-step real-time fluorescence mutant-allele-specific amplification （MASA） method for rapid SNP analysis. The method is a marriage of two technologies： MASA primers for target DNA and a double-stranded DNA-selective fluorescent dye, SYBR Green I. Genotypes are separated according to the different threshold cycles of the wild-type and mutant primers. K-ras oncogene was used as a target to validate the feasibility of the method. The experimental results showed that the different genotypes can be clearly discriminated by the assay. The real-time fluorescence MASA method will have an enormous potential for fast and reliable SNP analysis due to its simplicity and low cost.

Analysis of Single Nucleotide Polymorphism in Human Paraoxonase 1 Gene(Q192R) with Matrix-assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry

Introduction Single nucleotide polymorphisms （SNPs） are the most abundant DNA markers in the human genome occurring at a frequency of one in every 500--1000 nucleotides. A variety of methods have been used for the analysis of single nucleotide polymorphisms, including restriction fragment length polymorphism （RFLP）, direct sequencing by using laser-induced fluorescence detectionTM, fluorescence energy transfer, MALDI-TOF MS combined with primer extension or invasive cleavage, and fluorescence polarization. During the past two decades, mass spectrometry has become a very popular tool in the analysis of biomolecules and is perfectly suited to the analysis of single nucleotide polymorphisms （SNPs） due to its speed, low cost, and accuracy. In this work, we used MALDI TOF mass spectrometry to detect the fragments of restriction endonuclease hydrolysis of PCR products flanking a SNP located at paraoxonase 1（Q192R）. Compared with electrophoresis, this method requires less time of analysis and possess a higher accuracy.

Association between single nucleotide polymorphisms on chromosome 17q and the risk of prostate cancer in a Chinese population

In European populations,7 single nucleotide polymorphisms(SNPs) on chromosome 17q,3 SNPs on 17q12,and 4 SNPs on 17q24.3 were recently identified to be closely related to the risk of prostate cancer by a genome-wide association study.In Japanese populations,the correlation between 2 SNPs on 17q and the risk of prostate cancer and tumor aggressiveness was also confirmed by a large-scale experiment.However,whether 17q is associated with prostate cancer and its clinical manifestations in Chinese populations is still unknown.Therefore,we conducted a case-control study in a northern Chinese population and tested 2 SNPs,rs4430796 and rs1859962,on 17q in 124 prostate cancer patients and 111 controls using polymerase chain reaction-high resolution melting curve(PCR-HRM) combined with sequencing.We analyzed the association of the 2 SNPs with the risk of prostate cancer as well as patients' lifestyles,onset ages,Gleason scores,PSA levels,and pathologic stages.We found a significant difference in the G allele of SNP rs1859962(P = 0.035,OR = 1.51,95% CI = 1.03-2.21) but not in the rs4430796 genotype frequency or allele frequency distribution between prostate cancer patients and the controls(P > 0.05).Neither of the SNPs was significantly associated with the onset age,Gleason score,PSA level,pathologic stage,or other clinical indicators of patients with prostate cancer(P > 0.05).Our results show that polymorphism of the G allele of SNP rs1859962 is associated with the risk of prostate cancer in a Chinese population.