	US 11,987,822 B2
	Synthetic reverse transcriptases and uses thereof
Olev Kahre, Tartu (EE)
Assigned to SOLIS BIODYNE OÜ, Tartu (EE)
Filed by SOLIS BIODYNE OÜ, Tartu (EE)
Filed on May 25, 2021, as Appl. No. 17/329,358.
Application 17/329,358 is a continuation of application No. 16/092,753, granted, now 11,046,940, previously published as PCT/IB2017/052114, filed on Apr. 12, 2017.
Claims priority of provisional application 62/321,692, filed on Apr. 12, 2016.
Prior Publication US 2021/0388327 A1, Dec. 16, 2021
Int. Cl. C12N 9/12 (2006.01); C12N 9/22 (2006.01)

CPC C12N 9/1276 (2013.01) [C12N 9/22 (2013.01); C12Y 207/07049 (2013.01); C12Y 301/26004 (2013.01); C07K 2319/85 (2013.01)]

9 Claims

1. A method for synthesizing complementary DNA (cDNA), comprising:

(a) providing an RNA molecule template for cDNA synthesis,

(b) providing a primer to initiate cDNA synthesis from the RNA molecule, and

(c) synthesizing cDNA initiated by the primer from the RNA molecule template using a non-natural chimeric reverse transcriptase comprising:

(i) a first domain comprising viral reverse transcriptase domain, and

(ii) a second domain comprising a ribonuclease polypeptide having ribonuclease activity but not polymerase activity, wherein the ribonuclease polypeptide is an RNase polypeptide selected from the group consisting of: Pyrococcus furiosus RNase H, Pyrococcus horikoshi RNase H, Thermococcus litoralis RNase H II, Thermus thermophilus RNase H, and Escherichia coli RNase H;

wherein the non-natural protein retains at least 25% of its reverse transcriptase activity at 37° C. after incubation at a temperature of at least 60° C. for at least 10 minutes.