Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Reverse transcriptase polymerase chain reaction (RT-PCR)-based assays to detect occult neoplastic cells offer the highest sensitivity for the study of tumour dissemination and minimal residual disease. The detection of small numbers of tumour cells in a clinical sample may result in a redefinition of what constitutes residual disease and relapse, affecting future patient management. However, there remains disparity in the published data on the clinical value of RT-PCR for the detection of circulating tumour cells. This most likely reflects differences in the methods for sample preparation, RNA extraction, and cDNA synthesis among laboratories. Consequently the need for implementation of standard quality control measures is pressing in order to facilitate meaningful assessment of the methodology and it's clinical value. A 2-day workshop organized by the immunotherapy subgroup of the EORTC Melanoma Cooperative Group was held on this topic at the Ludwig Institute in Epalinges-sur-Lausanne, Switzerland in January 1996, with Stefan Carrel as the local host. Many pertinent issues were discussed in great detail, covering every step from sample handling to quality control. This workshop resulted in a concerted action leading to the preparation of laboratory guidelines, which are summarized in this review.
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PMID:Polymerase chain reaction detection of circulating tumour cells. EORTC Melanoma Cooperative Group, Immunotherapy Subgroup. 957 29

The synthesis of the triphosphates of 4'-thiouridine and 4'-thiocytidine, 4'-thioUTP (7; thioUTP) and 4'-thioCTP (8; thioCTP), and their utility for SELEX (systematic evolution of ligands by exponential enrichment) is described. The new nucleoside triphosphate (NTP) analogs 7 and 8 were prepared from appropriately protected 4'-thiouridine and -cytidine derivatives using the one-pot method reported by J. Ludwig and F. Eckstein [(1989) J. Org. Chem., 54, 631-635]. Because SELEX requires both in vitro transcription and reverse transcription, we examined the ability of 7 and 8 for SELEX by focusing on the two steps. Incorporation of 7 and 8 by T7 RNA polymerase to give 4'-thioRNA (thioRNA) proceeded well and was superior to those of the two sets of frequently used modified NTP analogs for SELEX (2'-NH2dUTP and 2'-NH2dCTP; 2'-FdUTP and 2'-FdCTP), when an adequate leader sequence of DNA template was selected. We revealed that a leader sequence of about +15 of DNA template is important for the effective incorporation of modified NTP analogs by T7 RNA polymerase. In addition, reverse transcription of the resulting thioRNA into the complementary DNA in the presence of 2'-deoxynucleoside triphosphates (dNTPs) also proceeded smoothly and precisely. The stability of the thioRNA toward RNase A was 50 times greater than that of the corresponding natural RNA. With these successful results in hand, we attempted the selection of thioRNA aptamers to human alpha-thrombin using thioUTP and thioCTP, and found a thioRNA aptamer with high binding affinity (K(d) = 4.7 nM).
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PMID:New NTP analogs: the synthesis of 4'-thioUTP and 4'-thioCTP and their utility for SELEX. 1591 69