Plant colonization assays using natural microbial barcodes
AssigneePivot Bio, Inc.
InventorsDouglas Higgins, Andrew Abel Prior
Life of this patent
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Abstract
The present disclosure is drawn to methods of utilizing nucleic acid barcodes and corresponding amplifying sites in cells in which the barcodes naturally occur. These barcodes and amplifying sites are reconfigured into a single nucleic acid cassette that provides for ease of use in tagging particular species, strains, or variants of cells, each with a different barcode. These barcodes can be used to track the colonization capabilities of the barcoded cells. The present disclosure further provides for assays that utilize natural barcodes to measure relative microbial colonization ability of a plant root system.
Claims18
An engineered bacterial cell, comprising:
- (a) a native barcode polynucleotide cassette inserted into a genomic locus of the cell, the cassette having first, second and third native nucleotide sequences oriented 5′ to 3′;
- wherein,
- (i) the first native nucleotide sequence serves as a forward primer binding site;
- (ii) the second native nucleotide sequence serves as a barcode;
- and
- (iii) the third native nucleotide sequence serves as a reverse primer binding site,
- wherein the first, second and third native nucleotide sequences comprised within the native barcode polynucleotide cassette are from the same species as the engineered bacterial cell into which they are being inserted;
- and
- wherein the genomic locus into which the native barcode polynucleotide cassette is inserted is at a heterologous location, as compared to the location in the genome of the first, second and third native nucleotide sequence.
Dependencies are inferred from the claim text.