ABSTRACT
Genome-wide association studies (GWAS) have identified single nucleotide polymorphisms (SNPs) associated with increased risk for colorectal cancer (CRC). A molecular understanding of the functional consequences of this genetic variation is complicated because most GWAS SNPs are located in non-coding regions. We used epigenomic information to identify H3K27Ac peaks in HCT116 colon cancer cells that harbor SNPs associated with an increased risk for CRC. Employing CRISPR/Cas9 nucleases, we deleted a CRC risk-associated H3K27Ac peak from HCT116 cells and observed large-scale changes in gene expression, resulting in decreased expression of many nearby genes. As a comparison, we showed that deletion of a robust H3K27Ac peak not associated with CRC had minimal effects on the transcriptome. Interestingly, although there is no H3K27Ac peak in HEK293 cells in the E7 region, deletion of this region in HEK293 cells decreased expression of several of the same genes that were downregulated in HCT116 cells, including the MYC oncogene. Accordingly, deletion of E7 causes changes in cell culture assays in HCT116 and HEK293 cells. In summary, we show that effects on the transcriptome upon deletion of a distal regulatory element cannot be predicted by the size or presence of an H3K27Ac peak.
Subject(s)
Gene Expression Regulation, Neoplastic , Histones/metabolism , Transcriptome , Acetylation , Cell Proliferation , Colorectal Neoplasms/genetics , Colorectal Neoplasms/metabolism , Enhancer Elements, Genetic , Genome-Wide Association Study , HCT116 Cells , HEK293 Cells , Humans , Papillomavirus E7 Proteins/genetics , Polymorphism, Single Nucleotide , Protein Processing, Post-Translational , Sequence DeletionABSTRACT
A novelly improved polymerase chian reaction and immunochromatography test (PCR-ICT) hybrid assay comprising traditional multiplex-nested PCR and ICT, (a lateral-flow device) was developed for direct detection of Legionella bacteria from environmental cooling tower samples. The partial 16S rDNA (specific for Legionella spp.) and dnaJ (specific for Legionella pneumophila) genes from Legionella chromosome were first specifically amplified by multiplex-nested PCR, respectively, followed by detection using ICT strip. Reading of results was based on presence or absence of the two test lines on the strips. Presence of test line 1 indicated existence of Legionella spp. specific 16S rDNA and identified Legionella spp. Presence of test line 2 further indicated existence of dnaJ and thus specifically identified L. pneumophila. In contrast, for non-Legionellae bacteria no test line formation was observed. Results of direct detection of Legionella bacteria and L. pneumophila from water tower specimens by this assay showed 100% sensitivity, and 96.6% and 100% specificity, respectively compared with traditional culture, biochemical and serological identification methods. The PCR-ICT hybrid assay does not require sophisticated equipment and was proved to be practically useful in rapid and direct Legionellae detection from environmental water samples.
Subject(s)
Immunologic Tests/methods , Legionella pneumophila/isolation & purification , Legionella/isolation & purification , Polymerase Chain Reaction/methods , Water Microbiology , Water Supply/analysis , Base Sequence , Colony Count, Microbial , Legionella/genetics , Legionella pneumophila/genetics , Sensitivity and SpecificityABSTRACT
This study was designed to investigate the electrical conductance of meridian in the obese during weight reduction. Ten obese including overweight (Body Mass Index, BMI > 26) and 30 healthy (non-overweight) people were recruited from Chung Yuan Christian University. The obese subjects were instructed to follow a weight reduction program that included diet control, exercise and oral intake of Prozac. A device, the design of which is based on the Ryodoraku theorem, was used to measure the electrical conductance of 12 meridians on both sides of the subjects. The results showed that: (1) the average coefficient of variation of the electrical conductances in 24 meridians of the obese group was statistically different from that of the healthy group (p < 0.05); (2) the average coefficient of variation of electrical conductance in the obese after weight reduction was significantly decreased than before the weight reduction program (p < 0.05); (3) the BMI and the electrical conductance of meridian was correlated in the obese (r = -0.77, p < 0.001) as well as in the healthy group (r = -0.92, p < 0.001). These findings suggest that electrical conductance of meridians can be a parameter to monitor weight, especially for obese people.