Browse
Submit Data
Databases
API
Help

Dataset Information

17 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Optimized PCR with sequence specific primers (PCR-SSP) for fast and efficient determination of Interleukin-6 Promoter -597/-572/-174Haplotypes.

ABSTRACT: BACKGROUND: Interleukin-6 (IL-6) promoter polymorphisms at positions -597(G-->A), -572(G-->C) and -174(G-->C) were shown to have a clinical impact on different major diseases. At present PCR-SSP protocols for IL-6 -597/-572/-174haplotyping are elaborate and require large amounts of genomic DNA. FINDINGS: We describe an improved typing technique requiring a decreased number of PCR-reactions and a reduced PCR-runtime due to optimized PCR-conditions. CONCLUSION: This enables a fast and efficient determination of IL-6 -597/-572/-174haplotypes in clinical diagnosis and further evaluation of IL-6 promoter polymorphisms in larger patient cohorts.

SUBMITTER: Muller-Steinhardt M

PROVIDER: S-EPMC2797812 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Optimized PCR with sequence specific primers (PCR-SSP) for fast and efficient determination of Interleukin-6 Promoter -597/-572/-174Haplotypes.

Müller-Steinhardt Michael M Schulte Friederike F Klüter Harald H Bugert Peter P

BMC research notes 20091210

<h4>Background</h4>Interleukin-6 (IL-6) promoter polymorphisms at positions -597(G-->A), -572(G-->C) and -174(G-->C) were shown to have a clinical impact on different major diseases. At present PCR-SSP protocols for IL-6 -597/-572/-174haplotyping are elaborate and require large amounts of genomic DNA.<h4>Findings</h4>We describe an improved typing technique requiring a decreased number of PCR-reactions and a reduced PCR-runtime due to optimized PCR-conditions.<h4>Conclusion</h4>This enables a fa ...[more]

PMID: 20003261

Similar Datasets

Interleukin-6 gene promoter-572 C allele may play a role in rate of disease progression in multiple sclerosis.

Project description:Multiple sclerosis (MS) is an inflammatory demyelinating disease affecting the central nervous system. Although the exact pathogenesis of MS is unknown, it is generally considered to be an autoimmune disease, with numerous genetic and environmental factors determining disease susceptibility and severity. One important mediator of immune responses and inflammation is interleukin-6 (IL-6). Previously, elevated levels of IL-6 in mononuclear cells in blood and in brain tissue from MS patients have been reported. Various polymorphisms in the promoter region of the IL6 gene have also been linked with IL-6 protein levels. In MS, several small studies have investigated whether two IL6 promoter polymorphisms (-597 G>A and -174 G>C) correlate with MS susceptibility, but with varying results. In the present study, we analyzed these polymorphisms, together with an additional polymorphism (-572 G>C) in 279 healthy controls and 509 patients with MS. We found no significant differences between MS patients and healthy controls for the different -597 or -174 IL6 promoter alleles or genotypes. There was a slight reduction in the percentage of individuals with MS who carried a C allele at position -572, although this was not significant after correction for multiple comparisons. Interestingly, however, the -572 C allele showed a significant correlation with the MS severity score, suggesting a possible role in disease progression.

| S-EPMC3497346 | biostudies-literature

GenoFrag: software to design primers optimized for whole genome scanning by long-range PCR amplification.

Project description:Genome sequence data can be used to analyze genome plasticity by whole genome PCR scanning. Small sized chromosomes can indeed be fully amplified by long-range PCR with a set of primers designed using a reference strain and applied to several other strains. Analysis of the resulting patterns can reveal the genome plasticity. To facilitate such analysis, we have developed GenoFrag, a software package for the design of primers optimized for whole genome scanning by long-range PCR. GenoFrag was developed for the analysis of Staphylococcus aureus genome plasticity by whole genome amplification in approximately 10 kb-long fragments. A set of primers was generated from the genome sequence of S.aureus N315, employed here as a reference strain. Two subsets of primers were successfully used to amplify two portions of the N315 chromosome. This experimental validation demonstrates that GenoFrag is a robust and reliable tool for primer design and that whole genome PCR scanning can be envisaged for the analysis of genome diversity in S.aureus, one of the major public health concerns worldwide.

| S-EPMC373259 | biostudies-literature

LOMA: a fast method to generate efficient tagged-random primers despite amplification bias of random PCR on pathogens.

Project description:BACKGROUND: Pathogen detection using DNA microarrays has the potential to become a fast and comprehensive diagnostics tool. However, since pathogen detection chips currently utilize random primers rather than specific primers for the RT-PCR step, bias inherent in random PCR amplification becomes a serious problem that causes large inaccuracies in hybridization signals. RESULTS: In this paper, we study how the efficiency of random PCR amplification affects hybridization signals. We describe a model that predicts the amplification efficiency of a given random primer on a target viral genome. The prediction allows us to filter false-negative probes of the genome that lie in regions of poor random PCR amplification and improves the accuracy of pathogen detection. Subsequently, we propose LOMA, an algorithm to generate random primers that have good amplification efficiency. Wet-lab validation showed that the generated random primers improve the amplification efficiency significantly. CONCLUSION: The blind use of a random primer with attached universal tag (random-tagged primer) in a PCR reaction on a pathogen sample may not lead to a successful amplification. Thus, the design of random-tagged primers is an important consideration when performing PCR.

| S-EPMC2553803 | biostudies-literature

easyPAC: A Tool for Fast Prediction, Testing and Reference Mapping of Degenerate PCR Primers from Alignments or Consensus Sequences

Project description:Video abstract A video abstract by the authors of this paper is available. video-abstract8870.mov The PCR-amplification of unknown homologous or paralogous genes generally relies on PCR primers predicted from multi sequence alignments. But increasing sequence divergence can induce the need to use degenerate primers which entails the problem of testing the characteristics, unwanted interactions and potential mispriming of degenerate primers. Here I introduce easyPAC, a new software for the prediction of degenerate primers from multi sequence alignments or single consensus sequences. As a major innovation, easyPAC allows to apply all customary primer test procedures to degenerate primer sequences including fast mapping to reference files. Thus, easyPAC simplifies and expedites the designing of specific degenerate primers enormously. Degenerate primers suggested by easyPAC were used in PCR amplification with subsequent de novo sequencing of TDRD1 exon 11 homologs from several representatives of the haplorrhine primate phylogeny. The results demonstrate the efficient performance of the suggested primers and therefore show that easyPAC can advance upcoming comparative genetic studies.

| S-EPMC3310402 | biostudies-literature

Gemi: PCR primers prediction from multiple alignments.

Project description:Designing primers and probes for polymerase chain reaction (PCR) is a preliminary and critical step that requires the identification of highly conserved regions in a given set of sequences. This task can be challenging if the targeted sequences display a high level of diversity, as frequently encountered in microbiologic studies. We developed Gemi, an automated, fast, and easy-to-use bioinformatics tool with a user-friendly interface to design primers and probes based on multiple aligned sequences. This tool can be used for the purpose of real-time and conventional PCR and can deal efficiently with large sets of sequences of a large size.

| S-EPMC3535827 | biostudies-literature

Primers and copper responsive promoter design and data of real-time RT-PCR assay in filamentous fungus Trichoderma reesei.

Project description:This data article contains data related to the research article entitled "Copper-mediated on-off control of gene expression in filamentous fungus Trichoderma reesei" (Wang et al., 2017) [1]. Four kinds of copper responsive promoters were designed. Quantitative PCR (qPCR) was performed to determine relative mRNA levels of red fluorescent protein gene (rfp) extracted from cells grown under different concentrations of CuSO4. Three deletion vectors were constructed by using a copper-mediated self-excision cassette instead of a xylose-mediated self-excision cassette (Zhang et al., 2016) [2] to knock out xyn1, one of the two major specific endo-β-1,4-xylanases (Rauscher et al., 2006) [3], xyr1, the key transcriptional activator of cellulolytic and xylanolytic genes (Lichius et al., 2015) [4], and ace3, a factor essential for cellulase production (Häkkinen et al., 2014) [5]. This data article reports the primers, vector construction, and qPCR assay.

| S-EPMC5694953 | biostudies-literature

-572 G/C single nucleotide polymorphism of interleukin-6 and sepsis predisposition in chronic renal disease.

Project description:Single nucleotide polymorphisms (SNPs) of interleukin (IL)-6 are associated with the development of chronic renal disease (CRD). Their impact for sepsis in the field of CRD was investigated. One control cohort of 115 patients with CRD without infection and another case cohort of 198 patients with CRD and sepsis were enrolled. Genotyping at the -174 (rs1800795) and -572 positions of IL-6 (rs1800796) was done by restriction fragment length polymorphism. Circulating IL-6 was measured by an enzyme immunoassay. The GG genotype of rs1800796 was more frequent among cases (78.3%) than controls (62.6%). No difference in the genotype frequencies of rs1800795 between cases and controls were found. Odds ratio for sepsis was 2.07 (95%CI 1.24-3.44, p = 0.005) with the GG genotype of rs1800796, which was confirmed by logistic regression analysis taking into consideration the presence of chronic comorbidities. All-cause mortality until day 28 was similar between patients with the GG genotype and the GC/CC genotypes of rs1800796, but death caused from cardiovascular events not-related with infection was more frequent with the GG genotype (14.6% vs 2.4%, p = 0.031). Circulating IL-6 was greater among patients of the GC/CC genotypes of rs1800796 and multiple organ dysfunction (p = 0.013). The GG genotype of rs1800796 predisposes to sepsis in CRD and to 28-day mortality by sepsis-unrelated cardiovascular phenomena.

| S-EPMC7102147 | biostudies-literature

Interleukin-6 (IL-6)-597 A/G (rs1800797) & -174 G/C (rs1800795) gene polymorphisms in type 2 diabetes.

Project description:BACKGROUND & OBJECTIVES: Diabetes is a metabolic pro-inflammatory disorder characterized by chronic hyperglycaemia and increased levels of circulating cytokines suggesting a causal role for inflammation in its aetiology. In order to decipher the role of interleukin-6 (IL-6) in type 2 diabetes mellitus (T2DM) we analyzed two promoter polymorphisms -597 A/G (rs1800797) and -174 G/C (rs1800795) in T2DM cases from north India, and in healthy controls. METHODS: DNA was isolated from venous blood samples of T2DM patients (n=213) and normal healthy controls (n=145). Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed after biochemical analysis. The genotypic and allelic frequency distributions were analyzed. RESULTS: The clinical/biochemical parameters of T2DM cases when compared to controls showed a significant difference. No significant association was observed with -597A/G polymorphism while, -174 G/C showed a highly significant association (P<0.001). In haplotypic analysis, combination of -597GFNx01/-174CFNx01 showed significant association (P=0.010). INTERPRETATION & CONCLUSIONS: Our data suggest that IL-6 gene polymorphisms play a prominent role in T2DM disease susceptibility in population from north India.

| S-EPMC4181162 | biostudies-literature

Production of recombineering substrates with standard-size PCR primers.

Project description:Recombineering is a powerful method for DNA manipulation. It has advantages over restriction endonuclease-based methods and is usually rapid. Typically, recombineering uses long PCR primers (c. 65 bases), each of which contains a small region of target homology (c. 45 bases). We have developed a simple, albeit somewhat less rapid, strategy to create recombineering substrates that can use primers of ? 35 bases for all steps. The regions of homology can be several hundred base pairs in length to (1) increase the chance of obtaining the desired clone and/or (2) allow coliphage-based recombineering in some non-Escherichia coli bacteria. The method uses cloning techniques to construct a template for the generation of the recombineering substrate. Because the template is made from cloned DNA segments, the segments (including those for the homology regions) can be readily changed. During construction of the template plasmid, potential background transformants arising from the vector without insert are significantly reduced by cloning each segment with two restriction endonucleases that produce noncompatible ends. We have used this method to change the bla gene of pACYC177 to aadA, to add the MCS-lacZ? region from pBBR1MCS to IncQ plasmid vectors, and to make an oriT(IncP) -aacC1 cassette and add it to a plasmid.

| S-EPMC3510332 | biostudies-literature

Development of a universal intimin antiserum and PCR primers.

Project description:Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) constitute a significant risk to human health worldwide. A hallmark of both pathogens is their ability to produce characteristic attaching-and-effacing (A/E) lesions in intestinal epithelial cells. Genes encoding A/E lesion formation map to a chromosomal pathogenicity island termed the locus of enterocyte effacement (LEE). Intimin, an LEE-encoded bacterial adhesion molecule, mediates the intimate bacterium-host cell interaction characteristic of A/E lesions. On the basis of characterization of the C-terminal 280-amino-acid cell binding domain of intimin (Int280(661-939)), four distinct Int280 types (types alpha, beta, gamma, and delta) have been identified. Importantly, Int280alpha and Int280beta antisera specifically recognized their respective intimin types. Using a conserved region of the intimin molecule (Int(388-667)) and primers synthesized to generate the recombinant Int(388-667), we have now generated universal intimin antiserum and PCR primers that are reactive with the different intimin types expressed by both human and animal A/E lesion-forming strains. Use of immunogold electron microscopy to visualize intimin on the surfaces of EPEC and EHEC strains revealed, in general, a uniform distribution on the bacterial cell surface. However, a filamentous staining pattern was observed with a few strains expressing intimin gamma. Cloning of the intimin eae gene from one such strain (strain ICC57) into strain CVD206, an EPEC strain which harbors a null deletion in eae, produced a uniform intimin staining pattern indicating that, if the filamentous staining pattern defines a filamentous form of intimin gamma, it is dependent upon the genetic background of the strain and is not a feature of the intimin molecule.

| S-EPMC85821 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data