US 12,275,983 B2
Processes for the production of oligonucleotides
Andreas Crameri, Stevenage (GB); Malcolm Leithhead Hill, Stevenage (GB); and David Graham Tew, Stevenage (GB)
Assigned to GlaxoSmithKline Intellectual Property Development Limited, Stevenage (GB)
Filed by GLAXOSMITHKLINE INTELLECTUAL PROPERTY DEVELOPMENT LIMITED, Stevenage (GB)
Filed on Mar. 30, 2020, as Appl. No. 16/833,724.
Application 16/833,724 is a continuation of application No. 15/643,535, filed on Jul. 7, 2017, granted, now 10,640,812.
Claims priority of application No. 1612011 (GB), filed on Jul. 11, 2016.
Prior Publication US 2020/0277659 A1, Sep. 3, 2020
Int. Cl. C12Q 1/6811 (2018.01); C07H 1/00 (2006.01); C07H 21/00 (2006.01); C12N 9/00 (2006.01); C12Q 1/6855 (2018.01); C12Q 1/686 (2018.01); G01N 30/02 (2006.01)
CPC C12Q 1/6811 (2013.01) [C07H 1/00 (2013.01); C07H 21/00 (2013.01); C12N 9/93 (2013.01); C12Q 1/6855 (2013.01); C12Q 1/686 (2013.01); C12Y 605/01001 (2013.01); C07K 2319/20 (2013.01); C12Q 2600/106 (2013.01); C12Q 2600/156 (2013.01); G01N 2030/027 (2013.01)] 20 Claims
 
1. A continuous or semi-continuous flow process for purifying a single-stranded oligonucleotide product from impurity oligonucleotide strands, wherein the single-stranded oligonucleotide product has at least one modified nucleotide residue, wherein the modification is chosen from: modification at the 2′ position of the sugar moiety of the nucleotide residue, modification of the nucleobase of the nucleotide residue, and modification of the backbone of the nucleotide residue, the process comprising:
a) providing a template oligonucleotide (I) complementary to the sequence of the single-stranded oligonucleotide product, said template having properties that allow it to be separated from the single-stranded oligonucleotide product;
b) providing a pool of oligonucleotides (II) comprising the single-stranded oligonucleotide product and impurity oligonucleotide strands;
c) contacting the template oligonucleotide (I) and the pool of oligonucleotides (II) in conditions to allow annealing, thereby producing (i) annealed template and impurity oligonucleotide strands and (ii) annealed template and single-stranded oligonucleotide product;
d) changing the conditions to separate any impurity oligonucleotide strands, comprising denaturing the annealed template and impurity oligonucleotide strands;
e) separating the impurity oligonucleotide strands from the annealed template and single-stranded oligonucleotide product;
f) changing the conditions to separate the single-stranded oligonucleotide product, comprising denaturing the annealed template and single-stranded oligonucleotide product;
g) separating the single-stranded oligonucleotide product from the template; and
h) recycling the template.