Thermostable reverse transcriptases and uses thereof
View Patent ↗The present invention is in the fields of molecular and cellular biology. The invention is generally related to reverse transcriptase enzymes and methods for the reverse transcription of nucleic acid molecules, especially messenger RNA molecules. Specifically, the invention relates to reverse transcriptase enzymes which have been mutated or modified to increase thermostability, decrease terminal deoxynucleotidyl transferase activity, and/or increase fidelity, and to methods of producing, amplifying or sequencing nucleic acid molecules (particularly cDNA molecules) using these reverse transcriptase enzymes or compositions. The invention also relates to nucleic acid molecules produced by these methods and to the use of such nucleic acid molecules to produce desired polypeptides. The invention also concerns kits comprising such enzymes or compositions.
1. A method for making one or more nucleic acid molecules comprising mixing one or more nucleic acid templates with one or more polypeptides having reverse transcriptase activity wherein the reverse transcriptase is an M-MLV reverse transcriptase comprising amino acids 34-710 of SEQ ID NO: 2 and incubating the mixture under conditions sufficient to synthesize one or more first nucleic acid molecules complementary to all or at least a portion of the one or more nucleic acid templates.
2. The method of claim 1 , wherein the one more nucleic acid templates are RNA.
3. The method of claim 2 , wherein the nucleic acid templates are mRNA.
4. The method of claim 1 , wherein the synthesized molecule is labeled.