Enhancers activate transcription in a distance-, orientation-, and position-independent manner, which makes them difficult to be identified. Self-transcribing active regulatory region sequencing (STARR-seq) measures the enhancer activity of millions of DNA fragments in parallel. Here we used STARR-seq to generate a quantitative global map of rice enhancers. Most enhancers were mapped within genes, especially at the 5' untranslated regions (5'UTR) and in coding sequences. Enhancers were also frequently mapped proximal to silent and lowly-expressed genes in transposable element (TE)-rich regions. Analysis of the epigenetic features of enhancers at their endogenous loci revealed that most enhancers do not co-localize with DNase I hypersensitive sites (DHSs) and lack the enhancer mark of histone modification H3K4me1. Clustering analysis of enhancers according to their epigenetic marks revealed that about 40% of identified enhancers carried one or more epigenetic marks. Repressive H3K27me3 was frequently enriched with positive marks, H3K4me3 and/or H3K27ac, which together label enhancers. Intergenic enhancers were also predicted based on the location of DHS regions relative to genes, which overlap poorly with STARR-seq enhancers. In summary, we quantitatively identified enhancers by functional analysis in the genome of rice, an important model plant. This work provides a valuable resource for further mechanistic studies in different biological contexts.
Acetylation ; Base Sequence ; Deoxyribonuclease I ; metabolism ; Enhancer Elements, Genetic ; Epigenesis, Genetic ; Genes, Plant ; Genomics ; methods ; Histone Code ; genetics ; Histones ; metabolism ; Models, Genetic ; Oryza ; genetics ; Promoter Regions, Genetic ; genetics ; Repetitive Sequences, Nucleic Acid ; genetics ; Sequence Analysis, DNA ; Transcription, Genetic

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe cases of human respiratory disease. The current outbreak of infection with this virus in South Korea, which began on May 20, 2015, has infected 186 patients and caused 36 deaths within 2 months. In this study, to investigate the viral pathogen causing acute respiratory infections, multiplex/RT-PCR was performed on were obtained from nucleic acid of the Middle East Respiratory Syndrome-negative subjects. Viruses and atypical bacteria were detected in 39 of 337 (11.6%). Frequent viruses were human rhinovirus (n=11, 3.3%), human metapneumovirus (n=9, 2.7%), parainfluenza (n=9, 2.7%) and adenovirus (n=4, 1.2%). Mycoplasma pneumonia (M. pneumonia) was detected in 1.8 % (n=6). Out of 9 human metapneumovirus (hMPV) positive samples, 6 samples were successfully sequenced using F gene. And M. pneumoniae was sequencing of a repetitive region of the P1 gene. Phylogenetic analysis revealed that hMPV clustered into A2b lineage (n=4), B2 lineage (n=2) and M. pneumoniae clustered into two genotypes: Type 1 (n=4), Type 2a (n=2).
Adenoviridae ; Bacteria ; Genotype ; Humans ; Korea ; Metapneumovirus ; Middle East Respiratory Syndrome Coronavirus ; Middle East* ; Paramyxoviridae Infections ; Pneumonia ; Pneumonia, Mycoplasma ; Repetitive Sequences, Nucleic Acid ; Respiratory Tract Infections ; Rhinovirus

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe cases of human respiratory disease. The current outbreak of infection with this virus in South Korea, which began on May 20, 2015, has infected 186 patients and caused 36 deaths within 2 months. In this study, to investigate the viral pathogen causing acute respiratory infections, multiplex/RT-PCR was performed on were obtained from nucleic acid of the Middle East Respiratory Syndrome-negative subjects. Viruses and atypical bacteria were detected in 39 of 337 (11.6%). Frequent viruses were human rhinovirus (n=11, 3.3%), human metapneumovirus (n=9, 2.7%), parainfluenza (n=9, 2.7%) and adenovirus (n=4, 1.2%). Mycoplasma pneumonia (M. pneumonia) was detected in 1.8 % (n=6). Out of 9 human metapneumovirus (hMPV) positive samples, 6 samples were successfully sequenced using F gene. And M. pneumoniae was sequencing of a repetitive region of the P1 gene. Phylogenetic analysis revealed that hMPV clustered into A2b lineage (n=4), B2 lineage (n=2) and M. pneumoniae clustered into two genotypes: Type 1 (n=4), Type 2a (n=2).
Adenoviridae ; Bacteria ; Genotype ; Humans ; Korea ; Metapneumovirus ; Middle East Respiratory Syndrome Coronavirus ; Middle East* ; Paramyxoviridae Infections ; Pneumonia ; Pneumonia, Mycoplasma ; Repetitive Sequences, Nucleic Acid ; Respiratory Tract Infections ; Rhinovirus

Trichomoniasis is the most common curable sexually-transmitted infection (STI) worldwide. There are few reports on the prevalence of Trichomonas vaginalis in Korea. The purpose of this study was to examine the prevalence of trichomoniasis by PCR in Guri city, Korea. All adult women who visited Hanyang University Guri Hospital for health screening within the National Health Care Service were invited to participate in the study, and 424 women were enrolled between March and June 2011. PCR was used to detect Trichomonas vaginalis using primers based on a repetitive sequence cloned from T. vaginalis (TV-E650). Fourteen women (3.3%) were found to have T. vaginalis. All were over 50, and they were significantly older on average than the 410 Trichomonas-negative women (mean ages 63.4 vs 55.3 years). It seems that T. vaginalis infection is not rare in women receiving health screening, especially among those over 50.
Adult ; Clone Cells ; Delivery of Health Care ; Female ; Humans ; Korea* ; Mass Screening* ; Polymerase Chain Reaction* ; Prevalence* ; Repetitive Sequences, Nucleic Acid ; Trichomonas vaginalis

Tuberculosis, caused by the bacterium Mycobacterium tuberculosis, remains one of the most serious global health problems. Molecular typing of M. tuberculosis has been used for various epidemiologic purposes as well as for clinical management. Currently, many techniques are available to type M. tuberculosis. Choosing the most appropriate technique in accordance with the existing laboratory conditions and the specific features of the geographic region is important. Insertion sequence IS6110-based restriction fragment length polymorphism (RFLP) analysis is considered the gold standard for the molecular epidemiologic investigations of tuberculosis. However, other polymerase chain reaction-based methods such as spacer oligonucleotide typing (spoligotyping), which detects 43 spacer sequence-interspersing direct repeats (DRs) in the genomic DR region; mycobacterial interspersed repetitive units–variable number tandem repeats, (MIRU-VNTR), which determines the number and size of tandem repetitive DNA sequences; repetitive-sequence-based PCR (rep-PCR), which provides high-throughput genotypic fingerprinting of multiple Mycobacterium species; and the recently developed genome-based whole genome sequencing methods demonstrate similar discriminatory power and greater convenience. This review focuses on techniques frequently used for the molecular typing of M. tuberculosis and discusses their general aspects and applications.
Base Sequence ; Dermatoglyphics ; Genome ; Global Health ; Methods* ; Molecular Typing ; Mycobacterium tuberculosis* ; Mycobacterium* ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Repetitive Sequences, Nucleic Acid ; Tandem Repeat Sequences ; Tuberculosis

PURPOSE: Spinocerebellar ataxias (SCAs) are progressive neurodegenerative disorders with diverse modes of inheritance. There are several subtypes of SCAs. SCA 8, SCA 12, and SCA 17 are the less common forms of SCAs with limited information available on their epidemiological profiles in Korea. The purpose of this study was to investigate the prevalence of SCA8, SCA12, and SCA17 in Korea. MATERIALS AND METHODS: Ninety-six unrelated Korean patients were enrolled and showed normal trinucleotide repeats through polymerase-chain reaction (PCR) for the genes ATXN1, ATXN2, ATXN3, CACNA1A , and ATXN7, which correspond to SCA1, SCA2, SCA3, SCA6, and SCA7, respectively. PCR products from patients were further analyzed by capillary electrophoresis using fluorescence labeled primers for the genes ATXN8OS, PPP2R2B, and TBP, which correspond to SCA8, SCA12, and SCA17. RESULTS: Three patients had 104, 97, and 75 abnormal expanded repeats in the ATXN8OS gene, the causative gene for SCA8. None of the patients exhibited abnormal repeats in SCA12 and SCA17. Normal trinucleotide repeat ranges of the cohort in this study were estimated to be 17-34 copies (average, 24+/-4 copies) for SCA8, 7-18 copies (average, 13+/-3 copies) for SCA12, and 26-43 copies (average, 35+/-2 copies) for SCA17. CONCLUSION: This study demonstrated that SCA8, SCA12, and SCA17 are rare in Korean patients with SCA, and further genetic studies are warranted to enhance the mutation detection rate in the Korean SCA population.
Asian Continental Ancestry Group ; Cohort Studies ; Electrophoresis, Capillary ; Fluorescence ; Humans ; Korea ; Neurodegenerative Diseases ; Polymerase Chain Reaction ; Prevalence ; Repetitive Sequences, Nucleic Acid* ; Spinocerebellar Ataxias* ; Trinucleotide Repeats ; Wills

This study aims to investigate the genetic characteristics of BZLF1 gene and its promoter Zp of the epidemic strains in children with primary Epstein-Barr virus (EBV)-associated diseases. Total DNA was extracted from the peripheral blood of 134 children with EBV-associated infectious mononucleosis (EBV-IM) and 32 children with EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH) who were admitted to Beijing Children's Hospital from 2006 to 2011. The EBNA3C, BZLF1, and Zp genes were amplified by PCR assay. Typing of EBV was performed according to the size of the amplification product of EBNA3C gene; the amplification products of BZLF1 and Zp genes were subjected to direct sequencing, and sequence analysis was performed using BioEdit 7. 0. 9. The results were as follows: (1) EBV-1 was present in 140 samples (97.2%, 140/144) and EBV-II in 4 samples (2.8%, 4/144). (2) Three BZLF1 genotypes and their 12 subtypes (including 6 newly found subtypes) were detected in this study; there were no significant differences in the frequencies of BZLF1-A and BZLF1-B between the children with EBV-IM and EBV-HLH (P = 0.083); BZLF1-A1 was the dominant genotype in children with EBV-associated diseases; t BZLF1-A mostly had three 29-bp repeats in the first intron of BZLF1 gene, and BZLF1-B mostly had 30-bp repeats (P = 0.000), with the number of repeats varying from 1 to 13. (3) Four Zp genotypes were detected in this study, including Zp-P, Zp-V3, Zp-V4, and Zp-V1; there were no significant differences in the frequencies of these Zp genotypes between children with EBV-IM and EBV-HLH (P = 0.272, 0.252, 1.0, and 1.0, respectively). (4) The linkage analysis of BZLF1 gene and its promoter Zp showed that BZLF1-A1 was highly associated with Zp-V3 (P = 0.000), while BZLF1-B4 with Zp-P (P = 0.000); EBV-I + BZLF1 A1 was highly associated with Zp-V3 (P = 0.000), while EBV-I+BZLF1-B4 with Zp-P (P = 0.000). The conclusions are as follows: (1) BZLF1-A1 is the dominant genotype in children with EBV-associated diseases; there are mostly 29-bp repeats in the first intron of BZLF1 gene for BZLF1-A genotype and 30-bp repeats for BZLF1-B genotype. (2) Zp-P and Zp-V3 are dominant Zp genotypes of EBV in children, which shared similar detection rates. (3) BZLF1-A1 is highly associated with Zp-V3, while BZLF1-B4 with Zp-P; EBV-I+BZLF1-A1 is highly associated with Zp-V3, while EBV-I+BZLF1-B4 with Zp-P.
Child ; Child, Preschool ; China ; epidemiology ; Epstein-Barr Virus Infections ; epidemiology ; virology ; Female ; Genotype ; Herpesvirus 4, Human ; genetics ; physiology ; Humans ; Infant ; Infant, Newborn ; Introns ; genetics ; Male ; Promoter Regions, Genetic ; genetics ; Repetitive Sequences, Nucleic Acid ; genetics ; Trans-Activators ; genetics

Murine leukemia virus (MLV)-based retroviral vectors is widely used for gene transfer and basic research, and production of high-titer retroviral vectors is very important. Here we report that expression of the Y-box binding protein 1 (YB-1) enhanced the production of infectious MLV vectors. YB-1 specifically increased the stability of viral genomic RNA in virus-producing cells, and thus increasing viral RNA levels in both producer cells and virion particles. The viral element responsive to YB-1 was mapped to the repeat sequence (R region) in MLV genomic RNA. These results identified YB-1 as a MLV mRNA stabilizer, which can be used for improving production of MLV vectors.
Base Sequence ; Gene Expression ; Genetic Engineering ; methods ; Genetic Vectors ; biosynthesis ; genetics ; Genome, Viral ; genetics ; HEK293 Cells ; Humans ; Leukemia Virus, Murine ; genetics ; physiology ; RNA Stability ; genetics ; RNA, Viral ; chemistry ; genetics ; Repetitive Sequences, Nucleic Acid ; genetics ; Untranslated Regions ; genetics ; Virion ; genetics ; physiology ; Y-Box-Binding Protein 1 ; genetics

OBJECTIVEBy testing the changes of telomere binding protein in malignant transformation BEAS-2B cells induced by coal tar pitch smoke extracts, to study the role of protection of telomeres 1 (POT1), telomeric repeat binding factor 1 (TRF1) and TRF2 in tumorgenesis that contact with coal tar pitch.

METHODSThe BEAS-2B cells were induced by coal tar pitch smoke extracts to form malignant transformation cell model in vitro. The gene expression levels of mRNA were assessed by real-time quantitative RT-PCR, the protein expression variations were determined by cell culture overslip of immunohistochemical methods.

RESULTSIn malignant transformation cells, the mRNA expression level (POT1: 0.63 ± 0.04, TRF1: 0.36 ± 0.01) and the protein expression level (POT1: 0.36 ± 0.05, TRF1: 0.09 ± 0.03) of POT1 and TRF1 was statistically significant decreased compared to that of BEAS-2B group (mRNA: POT1: 1.00 ± 0.04, TRF1: 1.01 ± 0.16; protein: POT1: 0.55 ± 0.07, TRF1: 0.27 ± 0.07) and DMSO group (mRNA: POT1: 0.89 ± 0.12, TRF1: 0.90 ± 0.08; protein: POT1: 0.55 ± 0.10, TRF1: 0.26 ± 0.04) (P < 0.05); mRNA expression level (1.45 ± 0.07) and the protein expression level (0.88 ± 0.06) of TRF2 was increased compared to that of BEAS-2B group (mRNA: 1.00 ± 0.07, protein: 0.48 ± 0.06) and DMSO group (mRNA: 1.00 ± 0.06, protein: 0.50 ± 0.06) (P < 0.05).

CONCLUSIONThe change of gene and protein expression level in POT1, TRF1, and TRF2 involved in the process that evolved into malignant transformation in bronchial epithelial cells BEAS-2B induced by coal tar pitch smoke extracts.

Cell Line ; Cell Transformation, Neoplastic ; metabolism ; Coal Tar ; toxicity ; Epithelial Cells ; cytology ; metabolism ; pathology ; Humans ; Repetitive Sequences, Nucleic Acid ; Telomere-Binding Proteins ; genetics ; metabolism

OBJECTIVETo study the structural shifts of gut flora in rats with acute alcoholic liver injury (AALI), and the effect of jianpi huoxue decoction (JPHXD) on the gut flora.

METHODSThirty-six Sprague-Dawley rats were randomly allocated to the control, AALI and JPHXD groups equally. The rats in the control group were given water and those in AALI and JPHXD groups were given ethanol by intragastric gavage for 5 days, while rats in the JPHXD group were administered JPHXD simultaneously. The blood and liver tissue were collected at the end of the experiment. The activities of serum alkaline aminotransferase (ALT), aspartate aminotransferase (AST), hepatic γ-glutamyltranspetidase (γ-GT) and hepatic triglyceride (TG) levels were determined. Plasma endotoxin level in the portal vein was measured. Pathological changes of liver tissues were determined by hematoxylin and eosin (HE) staining and oil red O staining. The total DNA of gut flora were extracted from fecal samples by Bead-beating method and determined by ERIC-PCR fingerprint method. The similarity cluster analysis and principal component analysis were performed to analyze the ERIC-PCR fingerprint respectively.

RESULTSIn the AALI group, the ratio of liver/body weight, activities of ALT, AST and hepatic γ-GT, amount of hepatic TG were elevated significantly compared with those in the control group (all P<0.01). JPHXD decreased the ratio, activities of ALT, AST, γ-GT and TG significantly compared with those in the AALI group (P<0.05 or P<0.01). HE and oil red O staining showed that fat deposited markedly in liver tissue, while JPHXD alleviated pathological changes markedly. Plasma LPS level in rat portal vein in the AALI group increased significantly (P<0.01), but it was decreased significantly in the JPHXD group (P<0.01). The cluster analysis and principal component analysis of ERIC-PCR fingerprint showed that gut flora in the AALI group changed markedly, and JPHXD could recover gut flora to some extent.

CONCLUSIONSThe structure of gut flora shifted markedly during acute alcoholic liver injury, JPHXD had prevention effect through the modification of gut flora.

Animals ; Azo Compounds ; metabolism ; Bacteria ; genetics ; Body Weight ; Cluster Analysis ; Consensus Sequence ; genetics ; DNA Fingerprinting ; methods ; DNA, Intergenic ; genetics ; Drugs, Chinese Herbal ; therapeutic use ; Freezing ; Gastrointestinal Tract ; microbiology ; pathology ; Liver ; microbiology ; pathology ; Liver Diseases, Alcoholic ; drug therapy ; microbiology ; pathology ; Organ Size ; Phylogeny ; Polymerase Chain Reaction ; methods ; Principal Component Analysis ; Rats ; Rats, Sprague-Dawley ; Repetitive Sequences, Nucleic Acid ; genetics ; Staining and Labeling