Compositions and Methods for RT-PCR

摘要

The present invention relates to compositions and methods having propylene glycol and DNA polymerase for facilitating the rapid and efficient amplification of nucleic acid molecules and the detection and quantitation of RNA molecules, and for increasing the detection sensitivity and reliability through generation of secure cDNA molecules prior to gene-specific primer dependent amplification. The reagent mixture comprises a ready to use reagent solution, wherein the solution comprises: (a) propylene glycol in a concentration between about 20% and about 50%; (b) a viral reverse transcriptase; and (c) at least one DNA polymerases, in a buffer suitable for use in a reverse transcription reaction, wherein the buffer comprises a co-factor metal ion and nucleoside triphosphates.

主权项

1. A reagent mixture comprising a ready to use reagent solution, wherein the solution comprises:
(a) propylene glycol in a concentration between about 25% and about 45%; (b) a viral reverse transcriptase in a concentration sufficient for use in a reverse transcription reaction without adding additional reverse transcriptase wherein the viral reverse transcriptase is selected from the group consisting of Avian Myeloblastosis Virus Reverse Transcriptase, Respiratory Syncytial Virus Reverse Transcriptase, Moloney Murine Leukemia Virus Reverse Transcriptase, Human Immunodeficiency Virus Reverse Transcriptase, Equine Infectious Anemia Virus Reverse Transcriptase, Rous-Associated Virus 2 Reverse Transcriptase, Avian Sarcoma Leukosis Virus Reverse Transcriptase, RNaseH (−) Reverse Transcriptase, SuperScript II Reverse Transcriptase, and ThermoScript Reverse Transcriptase; and (c) one or more DNA polymerases selected from the group consisting of Thermus Aquaticus, Thermus Thermophilus, Thermus Filiformis, Thermus Brockianus, Thermotoga Maritima, Thermotogo Neapolitana, Pyrococcus Furiosus, Pyrococcus Woesei, Vent, Deep Vent, Kodakaraensis, and mutants thereof, in a buffer suitable for use in a reverse transcription reaction, wherein the buffer comprises: a co-factor metal ion necessary for reverse transcriptase activity; and nucleoside triphosphates.