US 12,473,594 B2
Chemical tagging-based method for modified nucleoside sequencing, enrichment, and measurement
Guifang Jia, Beijing (CN); Ye Wang, Beijing (CN); and Shunqing Dong, Beijing (CN)
Assigned to PEKING UNIVERSITY, Beijing (CN)
Appl. No. 17/624,761
Filed by PEKING UNIVERSITY, Beijing (CN)
PCT Filed Jul. 1, 2020, PCT No. PCT/CN2020/099707
§ 371(c)(1), (2) Date Jan. 4, 2022,
PCT Pub. No. WO2021/000889, PCT Pub. Date Jan. 7, 2021.
Claims priority of application No. 201910601133.7 (CN), filed on Jul. 4, 2019.
Prior Publication US 2023/0332219 A1, Oct. 19, 2023
Int. Cl. C12Q 1/68 (2018.01); C07H 21/02 (2006.01); C12Q 1/6809 (2018.01); C12Q 1/6869 (2018.01); C12Q 1/6886 (2018.01)
CPC C12Q 1/6869 (2013.01) [C07H 21/02 (2013.01); C12Q 1/6809 (2013.01); C12Q 1/6886 (2013.01); C12Q 2600/106 (2013.01)] 18 Claims
 
1. A chemical tagging method for m6A-containing nucleic acid enrichment, comprising:
(1) treating a sample containing RNA and/or DNA with FTO or formaldehyde, wherein m6A is converted into hm6A under the presence of m6A in the RNA and/or DNA;
(2) adding a thiol compound, wherein the hm6A is reacted with the thiol compound to produce dm6A;
(3) adding MTSEA-biotin in a biotin tagging system to react with the dm6A to obtain biotinylated RNA and/or DNA; and
(4) enriching the biotinylated RNA and/or DNA by using streptavidin;
wherein the thiol compound has a structure of formula (I):
R—SH  (I),
wherein R is selected from the group consisting of C1-C6 alkyl, C1-C6 alkyl substituted by one or more substituents selected form the group consisting of hydroxyl, thiol and combinations thereof, C1-C6 alkyl phenyl, phenyl substituted by amino and C1-C6 ether group substituted by thiol.