Barcoding sequences for identification of gene expression
AssigneeMicrosoft Technology Licensing, LLC
InventorKris K. Ganjam
Life of this patent
Estimating the term…
Abstract
Gene expression can be identified by analyzing a DNA sequence. The DNA sequence can include a barcode sequence that corresponds to a particular gene. The barcode sequence can be produced during the expression of a gene by first adding a Homologous Directed Repair (HDR) template including the barcode sequence into the DNA sequence of the gene and then splicing the barcode sequence out of an RNA precursor during the expression of the gene. As the barcode sequence is made available from the RNA precursor, it can be added to the DNA strand using HDR. The resulting DNA strand can be sequenced and the sequence data can be analyzed to identify the barcode sequence within the DNA sequence, which provides an indicator of the expression of the gene in DNA rather than RNA.
Claims20
A method comprising:
- producing a first homology directed repair (HDR) template including at least a first splicing region and a barcode region, the first splicing region including a first sequence of nucleotides that is recognized by an enzyme to produce a cut in the first splicing region and the barcode region including a sequence of nucleotides that corresponds to a gene;
- inserting the first HDR template into a target site of the gene using HDR;
- splicing, using the enzyme, the first HDR template in at least the first splicing region to produce a second HDR template, the second HDR template including a sequence of nucleotides that includes a portion of the first splicing region and the barcode region;
- inserting the second HDR template into a double stranded polynucleotide using HDR;
- sequencing the additional double stranded polynucleotide to produce sequencing data;
- and
- determining that the gene has been expressed based at least partly on identifying the sequence of nucleotides of the barcode region in the sequencing data.
A system comprising:
- a gene including a double stranded polynucleotide having a target site;
- an enzyme configured to create a double strand break in the double stranded polynucleotide of the gene at a cut site in the target site;
- and
- a homology directed repair (HDR) template including at least a first splicing region and a barcode sequence corresponding to the gene;
- wherein the HDR template is inserted into the target site with HDR at the cut site after the enzyme creates a break at the cut site.
A system comprising:
- a gene;
- a double stranded polynucleotide including a target site;
- a homology directed repair (HDR) template including a barcode region having a sequence of nucleotides that corresponds to the gene;
- and
- an enzyme configured to create a double strand break in the double stranded polynucleotide at the target site;
- wherein the HDR template is inserted into the double stranded polynucleotide by HDR to produce a modified double stranded polynucleotide.
Dependencies are inferred from the claim text.