Brassica genomic assays
AssigneeE. I. Du Pont de Nemours and Company
InventorsMatthew Curtis Harmon, Nancy L. Henderson, Cathy Xiaoyan Zhong
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Abstract
Methods and compositions for detecting, identifying, and quantifying Brassica A genomic DNA are described. The methods are specific to the Brassica A genome and do not cross-react with other Brassica species, crops or weedy relatives that could contribute to contamination of a canola field.
Claims27
A method of detecting and quantifying the amount of Brassica A genomic DNA in a sample, the method comprising:
- a) specifically amplifying a genomic DNA fragment of the Brassica A genome, wherein the amplified DNA fragment comprises at least one of the nucleotide sequences of a genomic region selected from the group consisting of SEQ ID NOS:
- 25, 26, 27, 28, and 35;
- and
- b) detecting and quantifying the Brassica A genome from the amplified fragment of the Brassica A genome.
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A method of determining adventitious presence of a Brassica transgenic event in a sample, the method comprising:
- a) obtaining a sample suspected of containing a Brassica transgenic event;
- b) performing a Brassica A genome specific polymerase chain reaction, with a primer, wherein the primer binds to a genomic region of the Brassica A genome, the genomic region selected from the nucleotide sequence of at least one of SEQ ID NOS:
- 29, 30, or 31 from nucleotide position about 50 to about 400;
- c) determining the total amount of Brassica A genomic DNA in the sample;
- d) performing an event specific quantitative assay for the transgenic event to determine the amount of the transgenic event DNA in the sample;
- and
- e) comparing the amount of the transgenic event to the total amount of Brassica in the sample.
An amplicon comprising at least one of the nucleotide sequences of a genomic region selected from the group consisting of SEQ ID NOS:
- 25, 26, 27, 28, and 35 wherein the amplicon is not larger than 500 base pairs.
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A detection kit comprising oligonucleotides comprising a nucleotide sequence selected from the group consisting of SEQ ID NOS:
- 17, 18, 19, 20, 21, 22, 23, and 24 wherein the oligo is less than about 50 nucleotides and one or more reaction components to perform a quantitative reaction.
A method of establishing purity of a Brassica seed lot, the method comprising performing a polymerase chain reaction wherein the oligonucleotide primers or probes are capable of discriminating the Brassica A genome from the Brassica B and C genomes, wherein the oligonucleotide primers and/or probes bind to a target region of the Brassica A genome, the target region comprising a nucleotide sequence selected from the group consisting of SEQ ID NOS:
- 25, 26, 27, 28, and 35.
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A method of determining the presence and/or quantity of the Brassica A genome, the method comprising:
- a) specifically hybridizing DNA of the Brassica A genome with a probe, wherein the probe selectively binds to at least one of the nucleotide sequences of a genomic region selected from the group consisting of SEQ ID NOS:
- 25, 26, 27, 28, and 35, optionally under high stringency conditions, and;
- b) detecting the presence and/or quantity of the Brassica A genome.
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Dependencies are inferred from the claim text.