Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe here a general and simple procedure for cDNA library construction making use of in vitro amplification of cDNA by polymerase chain reaction (PCR). The first-strand cDNA is synthesized from total RNA with a primer EcoRI-(dT)17 and oligo(dG) tailed. An oligonucleotide, EcoRI-BamHI-(dC)13, is used to prime the second-strand synthesis by the thermostable DNA polymerase of Thermus aquaticus. The double-stranded cDNA is then amplified directly by PCR. A study of the effect of the elongation time on the PCR products showed that a long extension time is necessary to overcome the size heterogeneity of the cDNA population. Starting from 1 microgram of total brain RNA, the products obtained ranged from 200 to more than 2000 bp. The presence of the myelin basic protein cDNA sequence was determined. A lambda gt10 library containing 2 x 10(6) clones was established with the amplified cDNA. No sequences originating from rRNA were detected by Southern blot analysis. The ability to produce representative cDNA libraries from minute amounts of total RNA by this protocol should have many applications to studies of gene expression in small amounts of tissues or cells.
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PMID:cDNA library construction from small amounts of unfractionated RNA: association of cDNA synthesis with polymerase chain reaction amplification. 169 56

Epstein-Barr virus-specific CD4+ T cells could be involved in the pathogenesis of multiple sclerosis, provided they can gain entry to the intrathecal compartment. The authors have previously demonstrated that cerebrospinal fluid T cells from multiple sclerosis patients recognize autologous Epstein-Barr virus-transformed B cells. They now report that CD4+ T cells specific for the Epstein-Barr virus DNA polymerase peptide EBV 627-641 were present in the cerebrospinal fluid from one of two multiple sclerosis patients, and that a high proportion of these CD4+ T cells cross-recognized an immunodominant myelin basic protein peptide, MBP 85-99. In the observed patient, the proportion of EBV 627-641-specific CD4+ T cells seemed to exceed 1/10,000 in cerebrospinal fluid, compared to approximately 1/100,000 in blood. These findings prove that Epstein-Barr-virus specific CD4+ T cells can gain access to the intrathecal compartment, and suggest that Epstein-Barr virus-specific CD4+ T cells could target myelin basic protein in the central nervous system.
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PMID:Cerebrospinal fluid CD4+ T cells from a multiple sclerosis patient cross-recognize Epstein-Barr virus and myelin basic protein. 1538 50

Viral infections are thought to play an important role in the pathogenesis of multiple sclerosis potentially through molecular mimicry, but direct evidence from humans and animal models remains inadequate. Based on the fact that amino acid homology has been found between viral and host encephalitogenic protein, we designed four viral peptides (peptides of HBV polymerase protein, large T protein of JC virus, EB virus DNA polymerase and alkaline exonuclease of Human herpesvirus 6) with limited homology to myelin basic protein and explored their clinical, immunological and histological characteristics in Lewis rats. The immunization with JC virus peptide induced slight clinical signs of EAE in Lewis rats. Immunological examination indicated that rats immunized with JC virus peptide triggered T-cell cross-reactivity against MBP68-86, but failed to induce antibody cross-reactivity with MBP68-86. Histological staining exhibited the infiltration of inflammatory T cells and the activation of microglia in spinal cords of rats immunized with MBP68-86 and JC virus peptide. Other three peptides had negative findings in Lewis rats. These results suggested that molecular mimicry could be an important factor in the pathogenesis of EAE induced with JC virus peptide by expanding a population of reactive T cells that recognize MBP68-86 in Lewis rats inferring a possible pathogenesis for molecular mimicry in MS.
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PMID:Induction of experimental autoimmune encephalomyelitis in Lewis rats by a viral peptide with limited homology to myelin basic protein. 1761 6