US 12,454,712 B2
Method for large-scale synthesis of long-chain RNA and method for site-specific modification of long-chain RNA
Yong Hu, Shenzhen (CN); and Miaomiao Zhang, Shenzhen (CN)
Assigned to ShenZhen Rhegen Biotechnology Co., Ltd., Shenzhen (CN)
Filed by ShenZhen Rhegen Biotechnology Co., Ltd., Shenzhen (CN)
Filed on Jul. 18, 2022, as Appl. No. 17/813,290.
Application 17/813,290 is a continuation of application No. PCT/CN2020/136416, filed on Dec. 15, 2020.
Claims priority of application No. 202010068138.0 (CN), filed on Jan. 20, 2020.
Prior Publication US 2022/0372538 A1, Nov. 24, 2022
Int. Cl. C12P 19/34 (2006.01); C12N 9/00 (2006.01); C12N 9/16 (2006.01)
CPC C12P 19/34 (2013.01) [C12N 9/16 (2013.01); C12N 9/93 (2013.01); C12Y 301/03033 (2013.01)] 20 Claims
 
1. A method for large-scale synthesis of a long-chain RNA, comprising:
(a) obtaining a mixture comprising a plurality of short RNA fragments and a plurality of splint DNA fragments, wherein the plurality of short RNA fragments are ligated to form the long-chain RNA, and wherein each of the plurality of splint DNA fragments comprises a sequence that is complementary to 3′ end of one of the plurality of short RNA fragments and 5′ end of its downstream short RNA fragment; and
(b) adding a DNA ligase and an RNA ligase to the mixture, thereby obtaining the long-chain RNA,
wherein the method further comprises a step (a′) after (a) but prior to (b), wherein step (a′) comprises site-specific chemical modification of one or more short RNA fragments of the plurality of short RNA fragments.