Cloning multiple control sequences into chromosomes or into artificial centromeres
AssigneeChildren's Hospital Medical Center of Akron
InventorsRoger Lebo, James Malone
Life of this patent
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Abstract
Artificially synthesizing 29 homozygous cystic fibrosis core panel controls demonstrates placing multiple homozygous mutant sequences on the same single control DNA sequence to streamline quality control by minimizing extra control assays, time, and costly formatted test materials and testing all controls during every test. Any rare or unavailable reported DNA sequence can be PCR amplified using primer pairs synthesized with the designated mutation or variant sequence with paired adjacent upstream and downstream primers to amplify target sequences in total genomic DNA.
Claims14
A method of optimizing quality control in a genetic test assay, the method comprising the steps of:
- testing for the presence of a normal gene nucleotide sequence portion at a pre-selected gene locus;
- testing for the presence of at least a first mutant gene nucleotide sequence portion at the pre-selected gene locus;
- and
- testing for interference by at least a first homologous nucleotide sequence portion.
A control comprising:
- At least a first nucleotide sequence, wherein the at least a first nucleotide sequence includes at least a first homologous nucleotide sequence portion and wherein the at least a first nucleotide sequence lacks a sufficiently large segment of the gene nucleotide sequence at an at least a first pre-selected gene locus to preclude detection of the gene nucleotide sequence at the at least a first pre-selected gene locus by an assay, and
- Adapted for use in the assay, wherein the assay is for the detection of mutations in the gene nucleotide sequence at the at least a first pre-selected gene locus.
A control comprising:
- a first nucleotide sequence portion, wherein the first nucleotide sequence portion includes at least a first homologous nucleotide sequence portion having a sufficient length to adequately imitate corresponding normal or mutant nucleotide sequence portions found at a pre-selected gene locus being tested in an assay and containing at least a first distinct nucleotide species, wherein the at least a first distinct nucleotide species is not found in either the normal or mutant nucleotide sequence portions and wherein the at least a first distinct nucleotide species is suitably distinct as to provide a means for confirmation that the first homologous nucleotide sequence portion is not being detected in the assay by a normal or a mutant sequence primer used in the assay;
- and
- a second nucleotide sequence portion, wherein the second nucleotide sequence portion includes a sufficiently large segment of a first mutant gene nucleotide sequence portion found at the pre-selected gene locus so as to be detectable by the assay.
Dependencies are inferred from the claim text.