3' Untranslated Regions ; genetics ; 5' Untranslated Regions ; genetics ; Enterovirus B, Human ; genetics ; Genome, Viral ; genetics ; Humans ; Poly A ; genetics

OBJECTIVEIn order to fully understand hepatitis c virus (HCV) genotype 3b, 1a, 2b and 6a infection in China, We built HCV 5'-noncoding region (5'-NCR) of different genotypes and subtypes.

METHODSThe classification HCV into variable genotypes (subtypes) was carried on by programs A, B and C A. Using a combination of three restriction endonuclease BHH' (BsrB I, Hae II, Hinf I) digestions at the same time. The distinct genotypes were classified into 5 groups: genotype 1 (1a, 1b), 6a, 2 (2a, 2b), genotype 3 (3a, 3b), genotype4 (4a). B. With regard to genotype 1, we could distinguish subtype 1a from 1b using BstU I digestion. C. Using restriction endonuclease Hae III, genotype 2a, 2b, 3b, 4a, 6a are differentiated respectively.

RESULTS(1) HCV genotype 1a, 1b, 2a, 2b, 3a, 3b, 4a, 6a are fully discriminated by comparison with the genotypes regular samples. (2) Of the 93 patients, HCV genotype distribution in China was 66.67% for 1b, 18.28% for 2a, 3.23% for 1b/2b, 3b, 2b respectively. 2.15% for 2a/2b, 1b/2a respectively. 1.08% for 1a.

CONCLUSIONThis research indicated that adoption of HCV 5'-NCR A B C restriction endonuclease digestions techniques, might be sensitive and efficient to detect HCV and discriminate HCV genotype (subtypes) 1a to 6a.

5' Untranslated Regions ; chemistry ; DNA Restriction Enzymes ; Genotype ; Hepacivirus ; classification ; genetics ; RNA, Viral ; analysis

SINE-VNTR-Alu (SVA) elements are present in hominoid primates and are divided into 6 subfamilies (SVA-A to SVA-F) and active in the human population. Using a bioinformatic tool, 22 SVA element-associated genes are identified in the human genome. In an analysis of genomic structure, SVA elements are detected in the 5' untranslated region (UTR) of HGSNAT (SVA-B), MRGPRX3 (SVA-D), HYAL1 (SVA-F), TCHH (SVA-F), and ATXN2L (SVA-F) genes, while some elements are observed in the 3'UTR of SPICE1 (SVA-B), TDRKH (SVA-C), GOSR1 (SVA-D), BBS5 (SVA-D), NEK5 (SVA-D), ABHD2 (SVA-F), C1QTNF7 (SVA-F), ORC6L (SVA-F), TMEM69 (SVA-F), and CCDC137 (SVA-F) genes. They could contribute to exon extension or supplying poly A signals. LEPR (SVA-C), ALOX5 (SVA-D), PDS5B (SVA-D), and ABCA10 (SVA-F) genes also showed alternative transcripts by SVA exonization events. Dominant expression of HYAL1_SVA appeared in lung tissues, while HYAL1_noSVA showed ubiquitous expression in various human tissues. Expression of both transcripts (TDRKH_SVA and TDRKH_noSVA) of the TDRKH gene appeared to be ubiquitous. Taken together, these data suggest that SVA elements cause transcript isoforms that contribute to modulation of gene regulation in various human tissues.
3' Untranslated Regions ; 5' Untranslated Regions ; Exons ; Gene Expression Profiling ; Genome, Human ; Genomics ; Humans ; Lung ; Organ Specificity ; Poly A ; Primates ; Protein Isoforms

BACKGROUND: TT virus (TTV) infection is highly prevalent in the general population and in the patients infected with hepatitis B virus (HBV) or hepatitis C vius (HCV). The aim of the present study was to assess the positive rates of TTV DNA using different PCR primer sets in healthy and HBV or HCV-infected individuals in Korea. METHODS: TTV DNA was investigated in serum samples of 69 healthy individuals and 59 HBV-infected and 34 HCV-infected individuals by nested PCR assays using primers from N22 region, 5'-untranslated region (UTR), and 3' UTR of viral genome. RESULTS: TTV DNA was detected in 43% of total study populations using N22 primers, in 69% using 5' UTR primers and, in 64% using 3' UTR primers. No significant difference was observed in the positive rates of TTV DNA between healthy and HBV or HCV- infected individuals. CONCLUSION: The PCR assays for TTV DNA using 5' UTR primers and 3' UTR primers exhibited higher positive rates than that of the assay using N22 primers without any significant difference between healthy and HBV or HCV-infected individuals.
3' Untranslated Regions ; 5' Untranslated Regions ; DNA ; Genome, Viral ; Hepatitis B virus ; Hepatitis B* ; Hepatitis C ; Hepatitis* ; Humans ; Korea ; Polymerase Chain Reaction* ; Torque teno virus*

OBJECTIVEClone of sucrose synthase of Dendribium officinale and expression analysis, to provide the theory basis for research the relationship between polysaccharide synthesis of D. officinale and sucrose synthase activity.

METHODAccording to homologous sequence of sucrose synthase gene on GenBank, application the technology of RT-PCR and RACE, clone the full length of D. officinale. Target gene amplified with T vector was transformed into competent E. coli. BL21, IPTG induced expression, SDS-PAGE analysis.

RESULTA full length cDNA encoding sucrose synthase was isolated from the D. officinale, named DOSS1, the GenBank accession number is HQ856835, the cDNA is 2781 bp in length containing an open reading frame of 2424 bp encoding 807 amino acids with a predicted molecular mass of 92.3 x 10(3), the deduced amino acid sequence of D. officinale sucrose synthase shares 95% identity with Mokara yellow (AF530568); shares 90% identity with Oncidium goldiana (AF530567); shares more than 80% with other monocotyledonous plants.

CONCLUSIONCloned the sucrose synthase gene and induced an obvious band successfully.

3' Untranslated Regions ; genetics ; 5' Untranslated Regions ; genetics ; Cloning, Molecular ; Dendrobium ; enzymology ; genetics ; metabolism ; Escherichia coli ; genetics ; Gene Expression Regulation, Plant ; Glucosyltransferases ; genetics ; metabolism ; Phylogeny ; Polysaccharides ; biosynthesis

BACKGROUND: Osteoarthritis (OA) is a common disease characterized by degenerating joint cartilage in the knee, hip, and hand. A functional single nucleotide polymorphism (SNP) +104T/C; rs143383 in the 5' untranslated region of the growth differentiation factor 5 (GDF5) gene was recently associated with susceptibility to OA in the Japanese and Chinese populations. METHODS: To investigate whether this association is present in the Korean population, the frequency of the polymorphism was investigated in 276 patients with knee OA and 298 healthy subjects as controls. Polymorphism analysis was performed by amplifying the core promoter region of the GDF5 gene and digesting it with the BsiEI restriction enzyme. RESULTS: The frequency of the TT, CT, and CC genotypes was 54.3% (150/276), 41.7% (115/276), and 4.0% (11/276), respectively, in patients with OA, and 53.4% (159/298), 37.9% (113/298), and 8.7% (26/298), respectively, in healthy controls. No significant differences in genotypic or allelic frequencies of the +104T/C SNP of the GDF5 gene were observed between patients with OA and controls. Also, no significant differences in allelic and genotypic frequencies were found when the individuals were stratified by age and gender. CONCLUSIONS: The results suggest that the +104T/C; rs143383 GDF5 core promoter polymorphism is not a risk factor for OA in the Korean population.
5' Untranslated Regions ; Asian Continental Ancestry Group ; Cartilage ; Genotype ; Growth Differentiation Factor 5 ; Hand ; Hip ; Humans ; Joints ; Knee ; Osteoarthritis ; Polymorphism, Single Nucleotide ; Promoter Regions, Genetic ; Risk Factors

BACKGROUND: The completion of the human genome project means that a high-quality reference sequence of the gene-rich portion of the human genome is now available. However, the strong influence of ethnic, geographical, and other characteristics of study populations on the frequencies of different single-nucleotide polymorphisms (SNPs) makes it questionable whether foreign SNP data should be used in domestic studies. METHODS: Twenty-seven possible candidate genes of antiepileptic drug (AED) resistance were resequenced in a DNA pool from 200 healthy Koreans to identify SNPs and calculate their minor allele frequencies (MAFs). RESULTS: A total of 98 SNPs were present in 22 of the 27 genes: 28 were in the coding regions, 34 were in introns, 23 were in 5' near genes, 10 were in 5' untranslated regions, and 3 were in 3' near genes. CONCLUSIONS: The comparative analysis using the pooled DNA adopted in the present study was highly reliable in estimating MAFs and was compatible with the common disease/common variant hypothesis. The reported data on 98 publicly available SNPs of genes possibly associated with AED resistance that be useful to researchers with limited availability of domestic SNP data.
5' Untranslated Regions ; Clinical Coding ; DNA ; Drug Resistance ; Gene Frequency ; Genome, Human ; Human Genome Project ; Humans ; Introns ; Polymorphism, Single Nucleotide

OBJECTIVETo develop a new DNA chip with coloration, which can be used for rapid and economical detection of the genotyping of hepatitis C virus (HCV).

METHODSProbes and primers were designed according to the sequence of HCV 5' non-coding region (5' NCR) to fabricate DNA chip. Experimental group consisted of 60 positive serum samples and control group consisted of 20 negative serum samples. To obtain the aimed gene, then they were hybridized with DNA chip. Finally, the results showed in a nylon film. The results of DNA sequencing of samples were used as the control in double blind experimental.

RESULTSUsing DNA chip, HCV was detected in positive of all serum specimens of experimental group and negative in control group. The determination of HCV genotype by DNA chip showed corresponding rate of 96.7% with those by sequence assay.

CONCLUSIONIt showed higher specialty and sensitivity using DNA chip to detect the genotype of HCV. It would be valuable for the clinical genotyping of HCV

5' Untranslated Regions ; genetics ; Base Sequence ; Genotype ; Hepacivirus ; classification ; genetics ; Molecular Sequence Data ; Oligonucleotide Array Sequence Analysis ; Sequence Analysis, DNA

OBJECTIVETo clone and investigate the polymorphism of the 5'-untranslated region (5'-UTR) of the human ABO gene, in order to provide the basis for exploring the transcriptional regulation of the human ABO histo-blood group genes.

METHODSABO phenotypes of 30 unrelated healthy blood donors were determined by serological technique, their genotypes were analyzed by sequencing the exons 6 and 7 of the ABO gene. Nearly 5 kb of the 5'-UTR of ABO gene was obtained by PCR amplification and sequencing was performed bidirectionally. Haplotypes of samples with heterozygous sites in the 5'-UTR of ABO gene were separated and analyzed after cloning.

RESULTSTwenty polymorphic sites were identified in these samples where ABO genotypes were consistent with serological phenotypes. It included sixteen nucleotide sequence variations, one 8 bp deletion, one 6 bp deletion/insertion, one 36 bp insertion and one 43 bp repeats. Among them, 11 were novel polymorphic sites. Seven different haplotypes of 5'-UTR of ABO gene were defined to the cis/trans linkage of those mutations.

CONCLUSIONThere were polymorphisms in the 5'-UTR of ABO gene and the nucleotide sequences near the proximal promoter were conservative.

5' Untranslated Regions ; genetics ; ABO Blood-Group System ; genetics ; Cloning, Molecular ; Genotype ; Haplotypes ; Humans ; Polymorphism, Genetic ; Sequence Analysis, DNA

OBJECTIVETo investigate the roles of Domain I and Domain II of hepatitis C virus (HCV) 5' untranslated region (UTR) in the translation initiation activity of HCV 5'UTR in different host cell lines.

METHODSThe eukaryotic expression plasmid pCMVNCRLuc (pCN1), in which full-length HCV 5'UTR regulates firefly luciferase expression, was modified by deleting Domain I and the downstream single-stranded sequence (43 bp in total) from the UTR (pCNl-d2), Domain I with the downstream single-stranded sequence and Domain II (118 bp in total) from the UTR (pCNl-d3), or the total UTR (pCNl-d5). The modified plasmids were transfected via liposome into different cell lines with pRL-TK plasmid co-transfected as the normalization control. At 36 h after the transfection, the total cellular RNA was harvested for semi-quantitative RT-PCR, and the relative expression activities of luciferase were assayed with a dual luciferase reporter gene assay system. The translation initiation activities of the truncated HCV 5'UTRs in different translation systems were analyzed.

RESULTSDeletion of Domain I and the downstream single-stranded sequence caused no significant changes of the translational activity of HCV 5'UTR in Hela or C6 cells, but decreased the translational activity by 46% in L-02 cells and increased the translational activity by 46% in 293T cells. Deletion of both Domain I and Domain II resulted in decreased translational activity of HCV 5'UTR by 51% in HeLa cells, but increased the translational activity by 40% in L-02 cells, 60% in C6 cells and 135% in 293T cells.

CONCLUSIONSDomain I and Domain II of HCV 5'UTR perform cell type-specific roles in HCV IRES-driven translation initiation.

5' Untranslated Regions ; Genes, Reporter ; HeLa Cells ; Hepacivirus ; genetics ; Humans ; Luciferases ; Plasmids ; Protein Biosynthesis ; genetics ; RNA, Viral ; genetics ; Transfection