Processes for detecting or quantifying more than one nucleic acid in library
This invention provides novel compositions and processes for analyte detection, quantification and amplification. Nucleic acid arrays and libraries of analytes are usefully incorporated into such compositions and processes. Universal detection elements, signaling entities and the like are employed to detect and if necessary or desirable, to quantify analytes. Amplification of target analytes are also provided by the compositions and processes of this invention.
1. A process for detecting or quantifying more than one nucleic acid of interest in a library comprising the steps of:
a) providing:
(i) an array of fixed or immobilized nucleic acids identical in part or whole to said nucleic acids of interest;
(ii) a library of nucleic acid analytes isolated from a biological source, which library of nucleic acid analytes may contain the nucleic acids of interest sought to be detected or quantified, wherein each of said nucleic acids of interest comprise at least one inherent universal detection target (UDT);
(iii) a universal detection element (UDE) which generates a signal directly or indirectly; and
(iv) polymerizing means for synthesizing nucleic acid copies of said nucleic acid analytes;
b) synthesizing one or more first nucleic acid copies which are complementary to all or part of said nucleic acid analytes and synthesizing sequences which are complementary to all or part of said UDT to form a complementary UDT;
c) hybridizing said first nucleic acid copies with said array of nucleic acids (i) to form hybrids if said nucleic acids of interest are present;
d) contacting said UDEs with said complementary UDTs of said first nucleic acid copies to form a complex bound to said array; and
e) detecting or quantifying said more than one nucleic acid of interest by detecting or measuring the amount of signal generated from UDEs bound to said array.
2. The process of claim 1 , wherein said nucleic acid array comprises DNA, RNA or analogs thereof.
3. The process of claim 2 , wherein said nucleic acid array comprises analogs, wherein said analogs comprise PNA.
4. The process of claim 2 or 3 , wherein said DNA, RNA or analogs are modified on any one of the sugar, phosphate or base moieties.
5. The process of claim 1 , wherein said array comprises a solid support that is porous or non-porous.
6. The process of claim 5 , wherein said solid support is a porous solid support that is selected from the group consisting of polyacrylamide and agarose.
7. The process of claim 5 , wherein said solid support is a non-porous solid support that comprises glass or plastic.
8. The process of claim 5 , wherein said solid support is transparent, translucent, opaque or reflective.
9. The process of claim 1 , wherein said nucleic acids are indirectly fixed or immobilized to said solid support.
10. The process of claim 9 , wherein said nucleic acids are indirectly fixed or immobilized to said solid support by means of a chemical linker or linkage arm.
11. The process of claim 1 , wherein said biological source comprises an organ, a tissue or a cell.
12. The process of claim 1 , wherein said analytes are selected from the group consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and a combination of any of the foregoing.
13. The process of claim 1 , wherein said inherent UDT is selected from the group consisting of a poly T segment, a secondary structure, a consensus sequence, and a combination of any of the foregoing.
14. The process of claim 13 , wherein said inherent UDT is a consensus sequence selected from the group consisting of a signal sequence for polyA addition, a splicing element, a multicopy repeat, and a combination of any of the foregoing.
15. The process of claim 1 , wherein said UDE is selected from the group consisting of a nucleic acid, a nucleic acid analog, a polypeptide, a polysaccharide, a synthetic polymer and a combination of any of the foregoing.
16. The process of claim 15 , wherein said UDE is a nucleic acid analog that comprises PNA.
17. The process of claim 1 , wherein said UDE generates a signal directly.
18. The process of claim 17 , wherein said direct signal generation utilizes a fluorescent compound, a phosphorescent compound, a chemiluminescent compound, a chelating compound, an electron dense compound, a magnetic compound, an intercalating compound, an energy transfer compound or a combination of any of the foregoing.
19. The process of claim 1 , wherein said UDE generates a signal indirectly.
20. The process of claim 19 , wherein said indirect signal generation utilizes an antibody, an antigen, a hapten, a receptor, a hormone, a ligand, an enzyme or a combination of any of the foregoing.
21. The process of claim 20 , wherein said indirect signal generation utilizes an enzyme that catalyzes a reaction selected from the group consisting of a fluorogenic reaction, a chromogenic reaction and a chemiluminescent reaction.
22. The process of claim 1 , wherein said polymerizing means utilizes an enzyme selected from the group consisting of E. coli DNA Pol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reverse transcriptase, MuLV reverse transcriptase, RSV reverse transcriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscript and Omniscript.
23. A process for detecting or quantifying more than one nucleic acid of interest in a library comprising the steps of:
a) providing:
(i) an array of fixed or immobilized nucleic acids identical in part or whole to said nucleic acids of interest;
(ii) a library of nucleic acid analytes isolated from a biological source, which library of nucleic acid analytes may contain the nucleic acids of interest sought to be detected or quantified, wherein each of said nucleic acids of interest comprise at least one inherent universal detection target (UDT);
(iii) a universal detection element (UDE) which generates a signal directly or indirectly; and
(iv) polymerizing means for synthesizing nucleic acid copies of said nucleic acid analytes;
b) synthesizing one or more first nucleic acid copies of said nucleic acid analytes;
c) contacting said UDE with said UDT in said first nucleic acid copies to form one or more complexes;
d) hybridizing said first nucleic acid copies comprising said one or more complexes from said contacting step c) with said array of nucleic acids (i) to form hybrids if said nucleic acids of interest are present; and
e) detecting or quantifying said more than one nucleic acid of interest by detecting or measuring the amount of signal generated from any UDE bound to said array.
24. The process of claim 23 , wherein said nucleic acid array comprises DNA, RNA or analogs thereof.
25. The process of claim 24 , wherein said nucleic acid array comprises analogs, wherein said analogs comprise PNA.
26. The process of claim 24 or 25 , wherein said DNA, RNA or analogs are modified on any one of the sugar, phosphate or base moieties.
27. The process of claim 23 , wherein said array comprises a solid support that is porous or non-porous.
28. The process of claim 27 , wherein said solid support is a porous solid support that is selected from the group consisting of polyacrylamide and agarose.
29. The process of claim 27 , wherein said solid support is a non-porous solid support that comprises glass or plastic.
30. The process of claim 27 , wherein said solid support is transparent, translucent, opaque or reflective.
31. The process of claim 27 , wherein said nucleic acids are indirectly fixed or immobilized to said solid support.
32. The process of claim 31 , wherein said nucleic acids are indirectly fixed or immobilized to said solid support by means of a chemical linker or linkage arm.
33. The process of claim 23 , wherein said biological source comprises an organ, a tissue or a cell.
34. The process of claim 23 , wherein said analytes are selected from the group consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and a combination of any of the foregoing.
35. The process of claim 23 , wherein said inherent UDT is selected from the group consisting of a poly T segment, a secondary structure, a consensus sequence, and a combination of any of the foregoing.
36. The process of claim 35 , wherein said inherent UDT is a consensus sequence selected from the group consisting of a signal sequence for polyA addition, a splicing element, a multicopy repeat, and a combination of any of the foregoing.
37. The process of claim 23 , wherein said UDE is selected from the group consisting of a nucleic acid, a nucleic acid analog, a polypeptide, a polysaccharide, a synthetic polymer and a combination of any of the foregoing.
38. The process of claim 37 , wherein said UDE is a nucleic acid analog that comprises PNA.
39. The process of claim 23 , wherein said UDE generates a signal directly.
40. The process of claim 39 , wherein said direct signal generation utilizes a fluorescent compound, a phosphorescent compound, a chemiluminescent compound, a chelating compound, an electron dense compound, a magnetic compound, an intercalating compound, an energy transfer compound or a combination of any of the foregoing.
41. The process of claim 23 , wherein said UDE generates a signal indirectly.
42. The process of claim 41 , wherein said indirect signal generation utilizes an antibody, an antigen, a hapten, a receptor, a hormone, a ligand, an enzyme or a combination of any of the foregoing.
43. The process of claim 42 , wherein said indirect signal generation utilizes an enzyme that catalyzes a reaction selected from the group consisting of a fluorogenic reaction, a chromogenic reaction and a chemiluminescent reaction.
44. The process of claim 33 , wherein said polymerizing means utilizes an enzyme selected from the group consisting of E. coli DNA Pol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reverse transcriptase, MuLV reverse transcriptase, RSV reverse transcriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscript and Omniscript.
45. The process of claim 1 , wherein in said providing step a), the nucleic acid analytes are copies of nucleic acids from said biological source.
46. The process of claim 23 , wherein in said providing step a), the nucleic acid analytes are copies of nucleic acids from said biological source.
47. The process of claim 1 , wherein said nucleic acids are directly fixed or immobilized to said solid support.
48. The process of claim 27 , wherein said nucleic acids are directly fixed or immobilized to said solid support.