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US 2021/0032693 A1Published application
Improved Method to Analyze Nucleic Acid Contents from Multiple Biological Particles
AssigneeRootPath Genomics, Inc.
InventorsEly Porter, Xi Chen
Life of this patent
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Abstract
The application provides improved methods of analyzing biological particles and their constituents, including methods of labeling at least one target nucleic acid molecule from a biological particle with a barcoded primer.
Claims45
A method of labeling at least one target nucleic acid molecule from a biological particle with a barcoded primer, comprising:
- a. providing a pool of at least about 100 biological particles, wherein the biological particles comprise at least one target nucleic acid molecule;
- b. partitioning the pool of biological particles into compartments, wherein at least some of the compartments contain a primer delivery particle, wherein the primer delivery particle contains barcoded primers comprising at least 5 consecutive nucleotides that are complementary to at least a portion of the at least one target nucleic acid of the biological particle;
- and wherein the at least one barcoded primer binds to at least one target nucleic acid;
- and
- c. inactivating barcoded primers that are not bound to a target nucleic acid.
2 more elements
A method of labeling at least one target nucleic acid molecule from a biological particle with a barcoded primer, comprising:
- a. providing a pool of at least 100 biological particles, wherein the biological particles comprise at least one target nucleic acid;
- b. partitioning the pool of biological particles into compartments wherein at least some of compartments contain a primer delivery particle, wherein the primer delivery particle contains barcoded primers comprising at least 5 consecutive nucleotides that are complementary to at least a portion of at least one target nucleic acid of the biological particle;
- and wherein the at least one barcoded primer binds to at least one target nucleic acid;
- and
- c. mobilizing the barcoded primers from the primer delivery particles before and/or after the binding of at least one barcoded primer to at least one target nucleic acid;
- and
- d. heating the compartments accommodating the biological particles at a temperature of at least 80 degrees Celsius for at least 10 min
4 more elements
A method of labeling at least one target nucleic acid molecule from a biological particle with a barcoded primer, comprising:
- a. providing a pool of at least 100 biological particles, wherein the biological particles comprise at least one target nucleic acid;
- b. partitioning the pool of biological particles into compartments, wherein at least some of the compartments contain a primer delivery particle, wherein the primer delivery particle contains barcoded primers comprising at least 5 consecutive nucleotides that are complementary to at least a portion of at least one target nucleic acid of the biological particle;
- and wherein the at least one barcoded primer binds to at least one target nucleic acid;
- c. mobilizing the barcoded primers from the primer delivery particle before and/or after the binding of at least one barcoded primer to at least one target nucleic acid;
- and
- d. providing a fixation reversal agent in the compartments.
3 more elements
A method of labeling at least one target nucleic acid molecule from a biological particle with a barcoded primer, comprising:
- a. providing a pool of at least 100 biological particles, wherein the biological particles comprise at least one target nucleic acid;
- b. partitioning the pool of biological particles into compartments wherein at least some of the compartments contain a primer delivery particle, wherein the primer delivery particle contains barcoded primers comprising at least 5 consecutive nucleotides that are complementary to at least a portion of at least one target nucleic acid of the biological particle, and wherein the at least one barcoded primer binds to at least one target nucleic acid;
- and
- c. (i) mobilizing the barcoded primers from the primer delivery particle in the compartments before and/or after the binding of at least one barcoded primer to at least one target nucleic acid, (ii) after mobilizing the barcoded primers, pooling the contents of the compartments into an aqueous solution, and (iii) after pooling the contents, contacting the pooled contents in the aqueous solution with one or more nucleic acid polymerase.
1 more element
Dependencies are inferred from the claim text.