TaqI Restriction Site Research Articles

Molecular phylogenetic species in Alternaria pathogens infecting pistachio and wild relatives.

Many important pathogens of crops worldwide are members of section Alternaria within the genus Alternaria. Representative species in this section such as Alternaria alternata, Alternaria tenuissima, and Alternaria arborescens show high variability, intermediate characters and plasticity in morphological features, which makes species identification difficult. The aim of this study was to characterize Alternaria species associated with pistachio and wild relatives in Turkey using molecular phylogenetics. One hundred isolates of Alternaria spp. from pistachio and wild relatives from Turkey were investigated. In addition, standard morphological reference isolates and Alternaria blight pathogens of pistachio from USA were included. Sequence data from major allergen a1, ATPase, endopolygalacturanase, and anonymous regions OPA1.3 and SCAR2 were obtained. Gene trees were estimated based on maximum parsimony, maximum-likelihood, and bayesian inference methods. Species tree estimation was performed based on Yule speciation and strict molecular clock assumption. Among the collection of Alternaria spp. from Turkey, only one A. arborescens isolate and three isolates which were morphologically A. alternata/A. arborescens intermediate types, but, phylogenetically close to A. arborescens were discovered. While A. alternata and A. tenuissima formed one phylogenetic species, A. alternata/tenuissima were phylogenetically distinct from A. arborescens. Furthermore, a TaqI restriction site in the endopolygalacturanase gene was explored as a novel diagnosis for identification of A. alternata/tenuissima and A. arborescens. All these molecular phylogenetic approaches allow to distinguish morphologically similar Alternaria pathogens and molecular phylogenies of Alternaria pathogens from pistachio and wild relatives in Turkey are described for the first time.

Identification of a point mutation in the ace1 gene of Therioaphis trifolli maculata and detection of insecticide resistance by a diagnostic PCR-RFLP assay.

Aphids are important agricultural pests worldwide. Their control is largely based on chemical insecticides. One species that shows important invasive abilities and host-plant-related differences is Therioaphis trifolii (Monell) (Hemiptera: Aphididae). T. trifolii maculata, also known as spotted alfalfa aphid (SAA), can be very injurious to alfalfa crops in certain regions, such as in Saudi Arabia for effective control it is essential to diagnose and monitor the resistance mechanisms in the SAA populations. In the present study, we analysed acetylcholinesterase (ace) target site insensitivity mechanisms. A 650 bp length DNA containing the putative acetylcholinesterase (ace1) precursor was obtained and compared with other Hemipteran species. The sequences of many individual aphids collected from alfalfa crops in Saudi Arabia were analysed for the presence of resistance mutations: no resistance mutations were found at the resistance mutation loci 302; however, the presence of a serine-phenylalanine substitution (S431F) was identified in one individual. The S431F substitution, has been shown to confer significant levels of both organophosphate and carbamate resistance in other aphid species, and is now found for the first time in T. trifolii. We subsequently developed a simple polymerase chain reaction-restriction fragment length polymorphism assays for the S431F mutation, using a TaqI restriction site destroyed by the S431F mutation. The novel diagnostic assay may support the implementation of Insecticide Resistance Management strategies, for the control of SAA in alfalfa crops in the Kingdom of Saudi Arabia, and other countries worldwide.

DYZ1 arrays show sequence variation between the monozygotic males

BackgroundMonozygotic twins (MZT) are an important resource for genetical studies in the context of normal and diseased genomes. In the present study we used DYZ1, a satellite fraction present in the form of tandem arrays on the long arm of the human Y chromosome, as a tool to uncover sequence variations between the monozygotic males.ResultsWe detected copy number variation, frequent insertions and deletions within the sequences of DYZ1 arrays amongst all the three sets of twins used in the present study. MZT1b showed loss of 35 bp compared to that in 1a, whereas 2a showed loss of 31 bp compared to that in 2b. Similarly, 3b showed 10 bp insertion compared to that in 3a. MZT1a germline DNA showed loss of 5 bp and 1b blood DNA showed loss of 26 bp compared to that of 1a blood and 1b germline DNA, respectively. Of the 69 restriction sites detected in DYZ1 arrays, MboII, BsrI, TspEI and TaqI enzymes showed frequent loss and or gain amongst all the 3 pairs studied. MZT1 pair showed loss/gain of VspI, BsrDI, AgsI, PleI, TspDTI, TspEI, TfiI and TaqI restriction sites in both blood and germline DNA. All the three sets of MZT showed differences in the number of DYZ1 copies. FISH signals reflected somatic mosaicism of the DYZ1 copies across the cells.ConclusionsDYZ1 showed both sequence and copy number variation between the MZT males. Sequence variation was also noticed between germline and blood DNA samples of the same individual as we observed at least in one set of sample. The result suggests that DYZ1 faithfully records all the genetical changes occurring after the twining which may be ascribed to the environmental factors.

First Report of a 16SrIX Group ('Candidatus Phytoplasma phoenicium'-Related) Phytoplasma Associated with a Chrysanthemum Disease.

In November 2010, approximately 2% of the chrysanthemum (Chrysanthemum morifolium) cv. Paniz plants showed numerous small leaves in the top and stunting in a field collection of the National Research Center of Ornamental Plants in Mahallat, Iran. Next to these plants, some plants of the same collection showed leaves with a reddish and/or chlorotic discoloration around the veins. The observed symptoms were believed to represent infection by a phytoplasma and/or a viroid. Two plants with each type of the symptoms were individually analyzed. Using a total RNA extract from diseased leaves, RT-PCR with primer pairs targeting all known pospiviroids, including Chrysanthemum stunt viroid (CSVd) (3), were negative. Purified DNA was examined for the highly conserved phytoplasma 16S rRNA gene by nested-PCR using the universal primer sets P1/P7 and R16F2n/R16R2 (2). Fragments of 1.2 kb, obtained only from the plants with the small leaves and stunting, were sequenced and one of these sequences, which were identical, was deposited in GenBank (Accession No. KC176800). BLAST analysis of the chrysanthemum phytoplasma sequence exhibited 99% identity to Candidatus Phytoplasma phoenicium (Ca. P. phoenicium) species of the 16SrIX group. Subsequently, in silico RFLP analysis of the nested PCR product with the pDRAW32 program using AluI and TaqI restriction sites used for 16SrIX subgroups A, B, C, D, and E indicated that the 16SrIX chrysanthemum isolate belonged to subgroup D (1). Recently, based on GenBank sequences, several strains of Ca. P. phoenicium have been isolated and identified from diverse host species like Lactuca serriola, L. sativa, Solanum lycopersicon, Sonchus sp. [16SrIX-E], Carthamus tinctorius, and Prunus amygdalus [16SrIX-B] (4) in Iran. The vector species transmitting Ca. P. phoenicium to C. morifolium still needs to be identified. The leafhopper Neoaliturus fenestratus may be a potential vector as it is an often encountered efficient transmitter vector of 16SrIX group phytoplasmas in Iran (2). Next to the susceptibility of chrysanthemum to members of aster yellows, stolbur, and Ca. P. aurantifolia phytoplasma groups, this is, to our knowledge, the first report of a 16SrIX group member infecting chrysanthemum. The detection of this phytoplasma in chrysanthemum can form a new threat to this crop and other ornamentals in the Mahallat flower production region.

Function of TaGW2-6A and its effect on grain weight in wheat (Triticum aestivum L.)

Grain weight is an important factor contributing to grain yield in wheat. Markers closely linked to grain weight could be used in marker-assisted selection (MAS) to accelerate the wheat breeding process. In this paper, a second backcross recombinant inbred line (BC2F4) population, a recombinant inbred line (RIL) population, and one natural population were used to study the relationship between allelic variants of the grain weight gene TaGW2-6A and grain weight, width, and length with the goal of verifying the feasibility using the functional marker Hap-6A-P1/P2 in MAS. The differences in average grain weight, width, and length between haplotypes Hap-6A-G and Hap-6A-A were 8.09 g, 0.22 mm, and 0.38 mm, respectively, in the BC2F4 population; 4.01 g, 0.11 mm, and 0.10 mm in the RIL population; and 3.95 g, 0.10 mm, and 0.20 mm in the natural population. All the differences were significant. Sequencing results of the products of second round PCR indicated that a 167 bp TaqI restriction fragment had been generated from small-grain-size materials because of three TaqI restriction sites (TCGA). However, the large-grain-size variant generated a 218 bp fragment due to a single-nucleotide mutation (SNP) (TCGA → TCGG) at the third site. TaGW2-6A expression levels were negative with respect to grain width, length, and weight. For this reason, Hap-6A-G was considered a superior allele. These results contradict those of a previous study, which showed Hap-6A-A to be significantly associated with wider grain and higher grain weight.

Genetic Characterization of Insulin Growth Factor-1 and Its Receptor Genes in Egyptian Buffalo (Bubalus bubalis L.)

Aim: The somatotropic axis (SA) comprises genes associated with economically important quantitative traits in livestock like mammary and muscle growth as well as carcass traits. Insulin growth factor -1 (IGF-1) and its receptor ( IGF-1R) are two important genes belonging to the SA. The aim of this study was to evaluate the genetic polymorphism ofIGF1/SnaBI andIGF-1R/TaqIrestriction sites in Egyptian buffalo. Methodology :Genomic DNA was extracted from blood samples of 100 healthy buffaloes maintained at the Mahlet Mussa and ElGmeasa herds from 2010 to 2012. PCR was performed using primers flanking a 250 -bp fragment of the regulatory region of the buffalo IGF-1 gene and a616-bp fragment of theIGF-1R gene encompassing 51-bp from exon 12, 479-bp from intron 12 and 86bp from exon 13. The PCR-amplified fragments were digested withSnaBI (IGF-1) and TaqI (IGF-1R), electrophoresed and analyzed on agarose gels stained with ethi dium bromide.The two amplified fragments were also sequenced and aligned with published sequences. Results:All buffaloes investigated in this study were genotyped BB (i.e., negative for the SnaBI restriction site at position 224 ^225 (TAC^GTA) of theIGF-1regulatory region) and Research Article

Association Between Plasma 25-Hydroxyvitamin D and Colorectal Adenoma According to Dietary Calcium Intake and Vitamin D Receptor Polymorphism

The anticarcinogenic potential of vitamin D might be mediated by not only calcium metabolism but also other mechanisms initiated by vitamin D receptor (VDR). The authors measured plasma 25-hydroxyvitamin D in healthy volunteer examinees who underwent total colonoscopy in Tokyo, Japan, 2004-2005, and evaluated its influence on colorectal adenoma, both alone and in interaction with VDR polymorphisms, which correspond to the FokI and TaqI restriction sites. The main analysis of plasma 25-hydroxyvitamin D included 737 cases and 703 controls. Compared with the lowest quintile of plasma 25-hydroxyvitamin D, only the highest was related to a significantly decreased odds ratio of colorectal adenoma (odds ratio = 0.64, 95% confidence interval: 0.45, 0.92). In contrast, all but the lowest quintile of dietary calcium intake presented similarly reduced odds ratios (odds ratio for the highest = 0.67, 95% confidence interval: 0.47, 0.95). Of note, the association between plasma 25-hydroxyvitamin D levels and colorectal adenoma was modified by the TaqI polymorphism of the VDR gene (P(interaction) = 0.03) but not by dietary calcium intake (P(interaction) = 0.93). These observations highlight the importance of vitamin D in colorectal tumorigenesis. Vitamin D might protect against colorectal neoplasia, mainly through mechanisms other than the indirect mechanism via calcium metabolism.

Influence of vitamin D receptor haplotypes on blood lead concentrations in environmentally exposed children of Uygur and Han populations

Objective: To explore the association between polymorphism of vitamin D receptor (VDR) and lead poisoning in Uygur and Han children in China.Methods: The BsmI, ApaI and TaqI restriction sites of VDR genotyping were determined by polymerase chain reaction–restriction fragment length polymorphism in 443 Uygur and 469 Han children from Xinjiang province. The correlation between the polymorphism of VDR haplotypes and blood lead levels was explored.Results: The genotype frequencies of VDR had significant differences in Han and Uygur children (p <0.01). According to VDR–BsmI, ApaI and TaqI haplotype analysis in Han children, haplotypes Atb and AtB were considerably decreased in the lead poisoning group (p <0.05) while haplotype aTb and ATb were significantly increased in the lead poisoning group (p <0.01). However, such results were not found in Uygur children (p < 0.05).Conclusion: A significant difference was seen in the frequency distribution of the VDR genotype among the different races. Haplotypes Atb and AtB might be protective factors while haplotypes ATb and aTb might be risk factors in Han children.

Association between vitamin D receptor gene polymorphisms and iron indices in HIV-negative patients with malnutrition-inflammation-cachexia syndrome

Malnutrition-inflammation-cachexia syndrome (MICS) is a frequent complication of end-stage AIDS. Malnutrition, with its associated adverse effects on immunocompetence contributes to the progression of AIDS independently of HIV. Iron indices are considered reliable prognostic factors because of their association with inflammation and malnutrition. Since the active form of vitamin D has immunomodulatory effects, and considering that allelic variants of the vitamin D receptor (VDR) are associated with the rates of progression to AIDS in HIV-positive patients (J Steroid Biochem Mol Biol. 2004 89-90:199-207), here we evaluated the association between VDR polymorphisms and iron indices in HIV-negative patients with MICS. 38 HIV-negative patients treated at the Unit of Clinical and Artificial Nutrition, Misericordia e Dolce Hospital, Prato (Italy) for at least 12 months were studied. The mean age of the patients (22 men and 16 women) included in this study was 65 ± 10 years. Genomic DNA was extracted from peripheral blood leucocytes and amplified by polymerase chain reaction (PCR). PCR products were digested with the respective restriction enzymes in order to identify VDR polymorphisms. Absence or presence of the BsmI, ApaI, TaqI, and FokI restriction sites were denominated B and b, A and a, T and t, F and f respectively. Serum transferrin levels showed significant association with BsmI, ApaI and TaqI polymorphisms, i.e. those polymorphisms that are located in a regulatory site at the 3’ end of the VDR gene and are in linkage disequilibrium. Patients harbouring the BB, AA and tt genotypes showed significantly higher levels of serum transferrin compared with bb, Aa, aa, TT and Tt respectively. These results are consistent with those previously obtained in HIV-positive patients (J Steroid Biochem Mol Biol. 2004 89-90:199-207; J Infect Dis. 2008 197:405-10) and highlight an inverse correlation between vitamin D signalling and AIDS progression to MISC. These results also provide a link between VDR alleles and nutritional markers which are highly predictive variables of MICS. Since MICS is one of the leading causes of mortality in AIDS patients, the determination of VDR polymorphisms could help identifying those AIDS patients with a greater risk of developing MICS, a syndrome that appears to be independent of HIV serostatus or viral load.

Detection of Bovine Leukocyte Adhesion Deficiency (BLAD) in Turkish native and Holstein cattle

The purpose of this work was to study whether the bovine leukocyte adhesion deficiency (BLAD) allele is present in native cattle breeds and the Holstein breed in Turkey. Blood samples were obtained from 120 Holstein, 20 Brown Swiss, 20 Anatolian Black, 20 Turkish Grey, 20 South Anatolian Red and 20 East Anatolian Red cattle. The isolated DNA materials were multiplied in PCR using the primer developed by Kriegesmann et al. (1997). In order to determine the area of mutation in PCR products, the PCR products were digested with TaqI endonuclease enzyme. The resulting fragments were analysed on 2% agarose gel for the absence of a TaqI restriction site. It was found that two of the Holstein cattle (a bull and a cow) were heterozygote BLAD carriers. There was no homozygote BLAD animal. The BLAD allele was not found in the other breeds used in the study. The mutant BLAD allele frequency in the 120 Holstein cattle calculations was 0.0084.

Polymorphisms of vitamin D receptor gene protect against the risk of head and neck cancer

Vitamin D has potent anti-tumour properties. Calcitriol [1,25(OH)2D3], the hormonal derivative of vitamin D3, is an antiproliferative and prodifferentiation factor for several cell types, including human squamous cells of the head and neck. Several polymorphisms of the vitamin D receptor (VDR) gene have been described, including a FokI restriction fragment-length polymorphism (RFLP) in exon 2 and an adjacent TaqI RFLP in exon 9. We hypothesized that the VDR FokI and TaqI polymorphisms are associated with the risk of developing squamous cell carcinoma of the head and neck (SCCHN). We conducted a hospital-based, case-control study of 719 SCCHN cases and 821 cancer-free controls (all non-Hispanic Whites) to assess the association between VDR polymorphisms and SCCHN risk. The cases and controls were frequency-matched on age, sex and ethnicity. Polymorphisms at the TaqI and FokI restriction sites were determined from genomic DNA by polymerase chain reaction-RFLP methods. Both homozygous variant genotypes (ff and tt) were associated with a decreased risk of SCCHN [odds ratio (OR)=0.72, 95% confidence interval (CI)=0.53-0.98 and OR=0.64, 95% CI=0.47-0.87, respectively] compared to the common FF and TT genotypes. The VDR variant genotypes were associated with a decreasing risk of SCCHN in a variant allele dose-dependent manner, and the decreasing trend in OR was statistically significant, particularly for the combined genotypes (Ptrend<0.001). These data suggest that the VDR f and t alleles and their genotypes may protect against SCCHN. However, further studies are warranted to confirm these findings.

Risk of Early-Onset Prostate Cancer in Relation to Germ Line Polymorphisms of the Vitamin D Receptor

Abstract Vitamin D inhibits prostate cancer cell growth, angiogenesis, and metastasis. These actions are mediated by the vitamin D receptor. We examined associations between prostate cancer risk and five polymorphisms in the VDR gene: four single nucleotide polymorphisms (FokI, BsmI, ApaI, and TaqI restriction sites) and the polyadenylic acid microsatellite. Specifically, we genotyped population-based samples of young African Americans (113 cases and 121 controls) and Whites (232 cases and 171 controls) and members of 98 predominantly White families with multiple cases of prostate cancer. Among Whites, there was no evidence for association between prostate cancer risk and alleles at any of the five polymorphic sites regardless of how the men were ascertained. Moreover, estimated five-locus haplotype frequencies were similar in White cases and controls. Among African Americans, prostate cancer risk was associated with homozygosity for the F allele at the FokI site (odds ratio 1.9, 95% confidence interval 1.0-3.3). In addition, estimated haplotype frequencies differed significantly (P &amp;lt; 0.01) between African American cases and controls. These findings need replication in other studies of African Americans. Homozygosity for the F allele at the FokI site is more prevalent in the African American population than in U.S. Whites. If the FokI association noted here were causal, this difference could account for some of the disease burden among African Americans and some of the excess risk in African Americans compared with Whites.

Differentiation of infectious bursal disease viruses isolated in Croatia.

Infectious bursal disease viruses (IBDVs) in 26 IBDV-positive bursa samples collected in Croatia during the period 1996-2000 and in two commercially available vaccines were differentiated by the presence or absence of the CfoI, SacI, SspI, StuI, and TaqI restriction sites in the 422-bp fragment of segment A of the VP2 gene (nt 732-1153). The fragments from 14 (54%) field isolates were TaqI+ StuI+ SspI+ and SacI- CfoI-, indicating their very virulent (vv) character. The presence of CfoI restriction site in 10 (38%) field isolates is uncommon for vvIBDV strains. It was detected in only the 88180 vvIBDV strain. Nevertheless, these isolates can be classified as vv strains according to TaqI+ StuI+ SspI+ SacI- restrictions. Two SacI+ StuI+ CfoI+ TaqI- SspI- field isolates (8%) could be classified as non-vvIBDVs. The StuI+ restriction is common to vvIBDV strains. However, the StuI recognition sequence is present in the F52/70 classic European and 002-73 attenuated strains as well. The SacI+ CfoI+ StuI- SspI- restrictions and the lack of the TaqI restriction at nt position 832 show that the IBDV in GUMBOKAL IM-SPF vaccine corresponds to the attenuated and/or vaccine strains. The TaqI restriction at nt position 875 suggests that the IBDV in GUMBOKAL SPF vaccine could belong to the mild strains.

Association of the Vitamin D Receptor Genotype with Bone Metastases in Breast Cancer Patients

This study was designed in order to evaluate specific vitamin D receptor (VDR) genotypes as indicators of the likelihood of developing osseous metastases in breast cancer patients. Therefore, we determined polymorphisms of the VDR gene in a study group comprising 183 breast cancer patients. Specific fragments spanning over intron 8 and exon 9 of the VDR gene were amplified by polymerase chain reaction. The fragments were then incubated with each of the specific endonucleases ApaI, BsmI or TaqI, respectively. The VDR gene polymorphisms were detected by the presence or absence of the particular restriction site using agarose gel electrophoresis. Statistical analyses revealed a significant correlation between both the VDR gene polymorphisms indicated as AA (absence of the ApaI restriction site in both alleles) or TT (absence of the TaqI restriction site in both alleles), respectively, and the occurrence of bone metastases. Patients with the AA genotype have a 1.7-fold increased risk of developing bone metastases, whereas patients with the TT genotype have a 0.5-fold risk. Neither other genotypes nor allelic combinations displayed any further correlation with the clinical stage. The data suggest that the AA genotype of the VDR gene might be useful to identify breast cancer patients with a high probability of forming occult bone metastases who are considered to benefit from an adjuvant bone-protective therapy.

Significance of Vitamin D Receptor Gene Polymorphism for Risk and Disease Severity of Prostate Cancer and Benign Prostatic Hyperplasia in Japanese

Objective: Recent studies have demonstrated an association between vitamin D receptor (VDR) genotype and prostate cancer. Currently, there is a scarcity of data regarding the association of VDR genotype with benign prostatic hyperplasia (BPH). The purpose of this study was to investigate the TaqI VDR polymorphism in Japanese prostate cancer patients, Japanese BPH patients and Japanese controls in order to determine if an association exists between VDR genotype and the risk of developing prostate cancer and BPH as well as disease severity. Methods: 110 prostate cancer patients, 83 BPH patients and 90 male age-matched controls were genotyped for a previously described TaqI restriction fragment length polymorphism at codon 352 in exon 9 of the VDR gene. Products were digested into T allele or t allele according to the absence or presence of a TaqI restriction site with individuals being classified as TT, Tt or tt. Results: The frequency of the genotype tt was higher in the control group (6.7%) compared to patients with prostate cancer (1.8%) and BPH (3.6%) but this was not statistically significant. However, the frequency of the genotype TT was significantly higher among prostate cancer patients with locally advanced or metastatic disease (T3/ T4/N1/M1) compared to controls (p = 0.001). In addition, the genotype TT was significantly higher among prostate cancer patients with a high Gleason grade of tumor (grade 5) compared to controls (p = 0.0001). In addition, the genotype TT was statistically higher in BPH patients with high prostate volume (volume >50 cm³) compared to controls (p = 0.001). Conclusion: These data demonstrate that VDR genotype plays an important role in determining the risk of more advanced and aggressive prostate cancer as well as prostatic enlargement in Japanese men.

Vitamin D receptor gene polymorphism and prostate cancer risk.

1,25-dihydroxyvitamin D, the active form of vitamin D, exerts antiproliferative effect on prostatic cells, mediated through the vitamin D receptor. In a case-control study, we examined whether the vitamin D receptor (VDR) gene polymorphism in exon 9 could affect prostate cancer susceptibility. One hundred ninety newly diagnosed prostate cancer patients and 190 age-matched men with benign prostatic hyperplasia (BPH), in whom the presence of prostate cancer was excluded clinically or histologically, were recruited for this study. The VDR TaqI polymorphism was investigated by polymerase chain reaction (PCR) following restriction fragment length polymorphism using DNA from lymphocytes. Depending on the presence or absence of the TaqI restriction site at the third position of codon 352, patients were classified as TT, Tt, or tt. The frequency of the tt genotype was not significantly different between prostate cancer patients (18%) and controls (12%; P = 0.07). The odds ratio (OR), calculated relative to individuals with the TT genotype was 1.76 (95% confidence limit (CL) = 0.90-3.45). After stratification for Gleason score and prostate specific antigen levels in a case-case comparison (n = 190), no significant associations with the VDR genotypes were detectable either. In this case-control study of Austrian Caucasians, no statistically significant association of the VDR TaqI polymorphism and prostate cancer risk was found.

Further evidence for the association between attention-deficit/hyperactivity disorder and the dopamine-beta-hydroxylase gene.

Attention-deficit/hyperactivity disorder (ADHD) is a very common and heterogeneous psychiatric disorder of childhood with marked inattentive, hyperactive, and impulsive symptoms. The DBH gene, the locus that encodes the enzyme dopamine-beta-hydroxylase (DbetaH), seems to be an important candidate gene for association studies, since DbetaH catalyzes the conversion of dopamine to norepinephrine. The aim of this study was to test for association between the DBH gene and ADHD in a sample of 88 Brazilian nuclear families. Haplotype relative risk (HRR) analysis of the DBH TaqI restriction site polymorphism showed a preferential transmission of the TaqI A2 allele in our whole ADHD sample (chi(2)=3.61, one-tailed P=0.03). The significant effect of the A2 allele was stronger when only families with no ADHD parental diagnosis were considered (chi(2)=5.42, one-tailed P=0.01). Our results suggest a contribution of this gene to ADHD susceptibility, partially replicating previous findings that have demonstrated an association between the DBH TaqI A2 allele and ADHD.

Vitamin D receptor gene polymorphisms, bone mineral density and bone turnover: FokI genotype is related to postmenopausal bone mass

The relationship between vitamin D receptor (VDR) intragenic polymorphisms FokI, BsmI, ApaI and TaqI and bone mineral density (BMD) or biochemical markers of bone remodeling were investigated in 114 Czech postmenopausal women, on the average 62.5±8.9 years of age. Restriction fragment length polymorphisms in the VDR gene were assessed by PCR amplification and digestion with restriction enzymes FokI, BsmI, ApaI, and TaqI recognizing polymorphic sites in the VDR locus. Bone mineral density was measured at the lumbar spine and at the hip by dual-energy X-ray absorptiometry (DEXA, g/cm2). After adjusting for age and the body mass index (BMI), subjects with the ff genotype had 9.4 % lower BMD at the hip than those with the Ff genotype (p=0.0459, Tukey´s test). FF individuals had an intermediate BMD at the hip. A similar pattern of lower lumbar spine BMD was also found in ff individuals, but it did not reach statistical significance. There was no relationship between BsmI, ApaI and TaqI VDR polymorphisms and BMD at any skeletal site. Subjects with Aa (ApaI) genotypes had higher levels of propeptide of type I collagen (PICP) than homozygous AA (p=0.0459, Tukey´s test). In FokI, BsmI and TaqI restriction sites the biochemical markers of bone remodeling did not differ by genotype. In addition, no significant difference was observed in VDR genotypic distribution between osteoporotic women and non-osteoporotic controls in the study group. To conclude, the FokI genotype of the vitamin D receptor gene is related to bone mass at the hip in Czech postmenopausal women, whereas the importance of remaining VDR genotypes was not evident.

Association of vitamin D receptor gene polymorphism with renal cell carcinoma in Japanese.

Molecular epidemiologic studies have reported a relationship between 1alpha,25 dihydroxyvitamin D3 (1,25(OH)2D3) and the development and progression of malignant tumors. (1,25(OH)2D3) exerts its biological activity by binding the vitamin D receptor (VDR), while recent studies have demonstrated that VDR gene polymorphisms affect serum levels of (1,25(OH)2D3). Serum levels of (1,25(OH)2D3) are reported to be significantly lower in patients with renal cell carcinoma (RCC) compared to non-cancer control patients. The purpose of this study was to investigate the TaqI VDR polymorphism in Japanese RCC patients and non-cancer controls in order to determine if an association exists between VDR genotype and the risk of developing RCC as well as clinical risk factors. A total of 102 RCC patients and 204 controls were genotyped for a previously described TaqI restriction fragment length polymorphism (RFLP) of the VDR gene. Products were digested into T allele or the t allele according to the absence or presence of a TaqI restriction site. Individuals were classified as TT, Tt or tt. The genotype TT was statistically more frequent among RCC patients (80.4%) compared to controls (61.8%) (OR = 2.54; 95% CI, 1.44-4.46; p = 0.0006). In addition, the occurrence of the genotype TT was significantly higher in patients with rapid-growth-type group (92.1%) compared to slow-growth-type group (73.4%) (OR = 4.22; 95% CI, 1.15-15.53; p = 0.0175). These data demonstrate that VDR genotype plays an important role in determining the risk of developing more aggressive RCC in Japanese.

Specific Polymerase Chain Reaction Identification of Venturia nashicola Using Internally Transcribed Spacer Region in the Ribosomal DNA

ABSTRACT A technique based on the polymerase chain reaction (PCR) was developed for the identification of Venturia nashicola using nucleotide sequence information of the ribosomal DNA region. The complete internal transcribed spacer (ITS) region of V. nashicola strains and phylo-genetically related species was amplified with the two universal ITS1 and ITS4 primers, sequenced, and digested with five restriction enzymes. The alignment of nucleotide sequences and analyses of digestion patterns indicated constant polymorphisms between V. nashicola and related species at nucleotides 126 and 127, which overlapped a TaqI restriction site. An oligonucleotide primer named A126 was designed for identifying this variable region. A primer set (A126 and ITS4) that allowed the amplification of a 391-bp DNA fragment within the ITS region by PCR was specific to V. nashicola when it was checked against fungal genomic DNAs of related fungi. This primer set was a good candidate for a species-specific reagent in a procedure for identification of V. nashicola by PCR.