NORMALIZATION OF NUCLEIC ACID LIBRARIES

• US 20190292592A1
• Filed: 04/03/2019
• Published: 09/26/2019
• Est. Priority Date: 05/25/2016
• Status: Active Grant

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Cellular Research, Inc. (Becton, Dickinson & Co)
• Inventors: Shum, Eleen, Fu, Glenn, Betts, Craig
• Granted Patent: US 11,845,986 B2
• Time in Patent Office: Days
• CPC Class Codes: C12N 15/1065 Preparation or screening of tagged libraries, e.g. tagged microorganisms by STM-mutagenesis, tagged polynucleotides, gene tags
• CPC Class Codes: C12N 15/1093 General methods of preparing gene libraries, not provided for in other subgroups
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2537/159 Reduction of complexity, e.g. amplification of subsets, removing duplicated genomic regions
• CPC Class Codes: C12Q 2563/179 the label being a nucleic acid
• CPC Class Codes: G16B 25/00 ICT specially adapted for hybridisation; ICT specially adapted for gene or protein expression
• CPC Class Codes: G16B 25/10 Gene or protein expression profiling; Expression-ratio estimation or normalisation

Abstract

This disclosure provides methods and compositions for removing one or more high abundance species from a plurality of nucleic acid molecules. In some embodiments, the methods and compositions can be used for normalizing nucleic acid libraries. In some embodiments, molecular labels are used in conjunction with the methods and compositions disclosed herein to improve sequencing efficiency.

Claims

1-122. -122. (canceled)

123. A composition, comprising:

a plurality of first oligonucleotides comprising an affinity moiety; and

a plurality of capture molecules immobilized on one or more solid supports, wherein the capture molecules are configured to specifically bind to the affinity moiety.

124. The composition of claim 123, wherein the affinity moiety is a functional group selected from the group consisting of biotin, streptavidin, heparin, an aptamer, a click-chemistry moiety, digoxigenin, primary amine(s), carboxyl(s), hydroxyl(s), aldehyde(s), ketone(s), and any combination thereof.

125. The composition of claim 123, wherein the affinity moiety is biotin and the capture molecule is streptavidin.

126. The composition of claim 123, wherein the affinity moiety is streptavidin and the capture molecule is biotin.

127. The composition of claim 123, wherein the solid support is a bead.

128. The composition of claim 123, wherein each of the plurality of first oligonucleotides comprises a universal primer binding site.

129. The composition of claim 123, wherein each of the plurality of first oligonucleotides comprises a stochastic barcode.

130. The composition of claim 123, further comprising a plurality of second oligonucleotides configured to specifically hybridize to first oligonucleotides or complementary thereof

131. The composition of claim 130, wherein each of the plurality of second oligonucleotides comprises a universal primer binding site.

132. The composition of claim 130, wherein each of the plurality of second oligonucleotides comprises a stochastic barcode.

133. The composition of claim 123, comprising a target-specific oligonucleotide that specifically binds to a high abundance nucleic acid species.

134. The composition of claim 123, comprising one or more reagents for nucleic acid extension reactions.

135. The composition of claim 134, comprising a reverse transcriptase.

136. The composition of claim 123, comprising one or more reagents for PCR reactions.