Title: Multiplexed binary assembly and quantitative tracking of bacterial populations

Abstract

Compositions and methods for tracking combinations of mutations in populations by coupling DNA assembly and paired-end sequencing technology with high-throughput sequencing. This allows for an unlimited number of mutation sites in a population of microorganisms to be identified, and allows sites to be sequenced across several replicates. Unique identifiers (DNA barcodes) can be used to measure fitness data by sequencing of the barcodes multiplexed with the mutation sites with high-throughput short sequencing read technology, allowing selections to be rapidly performed on populations of known combinatorial genotypes.

Inventors:

Gill, Ryan T.; Zeitoun, Ramsey Ibrahim

Issue Date:

2020-10-06

Research Org.:

Univ. of Colorado, Denver, CO (United States)

Sponsoring Org.:

USDOE Office of Science (SC), Biological and Environmental Research (BER)

OSTI Identifier:

1771467

Patent Number(s):

10793851

Application Number:

15/749,540

Assignee:

The Regents of the University of Colorado, a body corporate (Denver, CO)

Patent Classifications (CPCs):

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS

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C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

C12N15/10 - Processes for the isolation, preparation or purification of DNA or RNA
C12N15/1065 - {Preparation or screening of tagged libraries, e.g. tagged microorganisms by STM-mutagenesis, tagged polynucleotides, gene tags}
C12N15/11 - DNA or RNA fragments

C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS

C12Q1/686 - Polymerase chain reaction [PCR]
C12Q2525/191 - incorporating an adaptor
C12Q2531/113 - PCR
C12Q2535/122 - Massive parallel sequencing

C - CHEMISTRY C40 - COMBINATORIAL TECHNOLOGY C40B - COMBINATORIAL CHEMISTRY

C40B40/06 - Libraries containing nucleotides or polynucleotides, or derivatives thereof
C40B50/06 - Biochemical methods, e.g. using enzymes or whole viable microorganisms

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DOE Contract Number:  

SC0008812

Resource Type:

Patent

Resource Relation:

Patent File Date: 08/04/2016

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES