Polony fluorescent in situ sequencing beads

1. An array comprising a plurality of beads, wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality, and wherein the plurality of beads is immobilized in a semi-solid medium to form an array.

2. The array of claim 1 , wherein the semi-solid medium is attached to a solid support.

3. The array of claim 2 , wherein the support is a microscope slide or a flow cell.

4. The array of claim 2 , wherein the semi-sold medium has top and bottom surfaces and the solid support is attached to the bottom surface.

5. The array of claim 1 , wherein the individual beads comprise two, three or four different populations of nucleic acid molecules attached thereto, which molecules have at least 50% sequence identity to each other.

6. The array of claim 1 , wherein at least 40% of the individual beads have populations of nucleic acid molecules attached thereto, wherein the nucleic acid molecules within each such population have at least 50% sequence identity to each other.

7. The array of claim 1 , wherein the beads are immobilized as a monolayer.

8. The array of claim 1 , wherein the semi-solid medium has x, y and z axes, and the plurality of beads is randomly arranged relative to the x and y axes.

9. The array of claim 1 , wherein the semi-sold medium has top and bottom surfaces and the plurality of beads is immobilized near the top surface.

10. The array of claim 1 , wherein the semi-solid medium is selected from the group consisting of: polyacrylamide, cellulose, polyamide, cross-linked agarose, cross-linked dextran and cross-linked polyethylene glycol.

11. The array of claim 1 , wherein the plurality of beads comprises a plurality of clonal beads.

12. The array of claim 1 , wherein the plurality of beads comprises a library.

13. The array of claim 1 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

14. The array of claim 1 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

15. The array of claim 1 , wherein each of two, three or four individual beads has a population of nucleic acid molecules attached thereto having at least 50% sequence identity to each other.

16. The array of claim 15 , wherein each population of nucleic acid molecules attached to each of the two, three or four individual beads differs in sequence from one another.

17. The array of claim 15 , wherein each population of nucleic acid molecules attached to each of the two, three or four individual beads is identical in sequence to one another.

18. A method of producing an array comprising the steps of:

a) providing a plurality of beads wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality; and

b) immobilizing the beads in a semi-solid medium to form an array.

19. The method of claim 18 , further comprising attaching the semi-solid medium to a solid support in step b).

20. The method of claim 19 , wherein the support is a microscope slide or a flow cell.

21. The method of claim 19 , wherein the semi-sold medium has top and bottom surfaces and the solid support is attached to the bottom surface.

22. The method of claim 18 , wherein the plurality of beads comprises two, three or four different populations of nucleic acid molecules attached thereto, which molecules have at least 50% sequence identity to each other.

23. The method of claim 18 , wherein at least 40% of the individual beads have populations of nucleic acid molecules attached thereto, wherein the nucleic acid molecules within each population have at least 50% sequence identity to each other.

24. The method of claim 18 , wherein the beads are immobilized as a monolayer.

25. The method of claim 18 , wherein the semi-solid medium has x, y and z axes, and the plurality of beads is randomly arranged relative to the x and y axes.

26. The method of claim 18 , wherein the semi-sold medium has top and bottom surfaces and the plurality of beads is immobilized near the top surface.

27. The method of claim 18 , wherein the semi-solid medium is selected from the group consisting of: polyacrylamide, cellulose, polyamide, cross-linked agarose, cross-linked dextran and cross-linked polyethylene glycol.

28. The method of claim 18 , wherein the plurality of beads comprises a plurality of clonal beads.

29. The method of claim 18 , wherein the plurality of beads comprises a library.

30. The method of claim 18 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

31. The method of claim 18 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

32. The method of claim 18 , wherein each of two, three or four individual beads has a population of nucleic acid molecules attached thereto having at least 50% sequence identity to each other.

33. The method of claim 32 , wherein each population of nucleic acid molecules attached to each of the two, three or four individual beads differs in sequence from one another.

34. The method of claim 32 , wherein each population of nucleic acid molecules attached to each of the two, three or four individual beads is identical in sequence to one another.

35. A method of producing an array comprising the steps of:

a) providing a plurality of beads wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality;

b) immobilizing the beads in a semi-solid medium to form an array; and

c) amplifying the populations of nucleic acid molecules having at least 50% sequence identity to each other to form a plurality of beads having amplified populations of nucleic acid molecules having at least 50% sequence identity to each other attached thereto.

36. The method of claim 35 , wherein the semi-solid medium includes an amplification primer.

37. The method of claim 35 , wherein the semi-solid medium includes an additive that forms voids in the semi-solid medium.

38. The method of claim 37 , wherein the additive is selected from the group consisting of: cationic lipid, polyamine and polycation.

39. The method of claim 35 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

40. The method of claim 35 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

41. A method of producing an array comprising the steps of:

a) providing a plurality of beads wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality;

b) amplifying the populations of nucleic acid molecules having at least 50% sequence identity to each other to form a plurality of immobilized beads having amplified populations of nucleic acid molecules having at least 50% sequence identity to each other attached thereto; and

c) immobilizing the beads in a semi-solid medium to form an array.

42. The method of claim 41 , wherein the amplifying is performed by emulsion PCR.

43. The method of claim 41 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

44. The method of claim 41 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

45. A method of producing an array comprising the steps of:

a) providing a plurality of beads wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality;

b) amplifying the populations of nucleic acid molecules having at least 50% sequence identity to each other to form a plurality of beads having amplified populations of nucleic acid molecules having at least 50% sequence identity to each other attached thereto;

c) enriching the plurality of beads having amplified populations of nucleic acid molecules having at least 50% sequence identity to each other attached thereto to form an enriched population of beads; and

d) immobilizing the beads in a semi-solid medium to form an array.

46. The method of claim 45 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

47. The method of claim 45 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

48. A method for enriching a population of beads having a first nucleic acid molecule attached thereto comprising the steps of:

a) providing a population of beads wherein an individual bead has a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality;

b) contacting the population of beads with a second nucleic acid molecule immobilized on a support, wherein the second nucleic acid molecule has a sequence that is complementary to that of the first nucleic acid molecule;

c) incubating the population of beads and the second nucleic acid molecule together such that hybridization occurs to form a population of hybridized beads and a population of unhybridized beads; and

d) separating the population of hybridized beads from the population unhybridized beads.

49. The method of claim 48 , wherein the second nucleic acid is immobilized on a capture bead.

50. The method of claim 48 , wherein the population of hybridized beads are separated from the population of unhybridized beads by density or affinity.

51. The method of claim 48 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are primers.

52. The method of claim 48 , wherein the nucleic acid molecules having at least 50% sequence identity to each other are amplified nucleic acid molecules.

53. A kit containing an array comprising a plurality of beads immobilized in a semi-solid medium, wherein the plurality of beads includes an individual bead having a population of nucleic acid molecules having at least 50% sequence identity to each other attached thereto, which molecules differ in sequence from those of a population of nucleic acid sequences molecules having at least 50% sequence identity to each other attached to another individual bead included in such plurality.