초록 ▼

A new method has been developed for conducting a gene probe assay. The preferred technique involves (1) using a gene amplification technique (e.g., PCR) to multiply the gene sequence of interest and (2) using a hybridization--ligation detection methodology, wherein the sequences of probes hybridized

A new method has been developed for conducting a gene probe assay. The preferred technique involves (1) using a gene amplification technique (e.g., PCR) to multiply the gene sequence of interest and (2) using a hybridization--ligation detection methodology, wherein the sequences of probes hybridized to the target sequence allow for separation and detection (e.g., probes might contain a combination of magnetic particles and acridinium esters) to determine if a specific sequence is present.

대표청구항 ▼

[ What is claimed is:] [1.] A method for identifying a target polynucleic acid sequence in a reaction mixture, by detecting whether or not a label is present comprising:a) selecting a first probe and a second probe, such thati) they are each complementary to a sufficient portion of the target to per

[ What is claimed is:] [1.] A method for identifying a target polynucleic acid sequence in a reaction mixture, by detecting whether or not a label is present comprising:a) selecting a first probe and a second probe, such thati) they are each complementary to a sufficient portion of the target to permit the formation of a hybrid,ii) the first probe is joined to a moiety which is a means for selectively separating the first probe from the reaction mixture,iii) the second probe is joined to a label or means for selectively attaching a label,b) mixing the probes with the target polynucleic acid in such a manner that the probes will hybridize under high stringency conditions specifically to the target polynucleic acid,c) adding a ligating reagent,d) denaturing the hybridized probes and target polynucleic acid in the reaction mixture so that the probes will be separated from the target polynucleic acid,e) separating the first probe from the reaction mixture, utilizing the moiety that permits selective separation, andf) analyzing the selectively separated first probe to determine if the label, or means for attaching a label, of the second probe is present, whereby the identification of the target polynucleotide sequence is made, by correlating the presence or absence of label or means for attaching the label on the second probe with the presence or absence of the target polynucleotide, and analyzing the supernatant remaining after separation of the first probe to determine the amount of label present therein and, further, to determine the percent of hybridized labeled probe which has been ligated to said first probe,wherein a step for separating the hybridized probes and target, utilizing the moiety that permits selective separation, must take place either (1) before ligating step c), (2) after ligating step c), or (3) both before and after ligating step c).