Polycationic supports for nucleic acid purification, separation and hybridization

Claim 2

A method according to claim 1 further comprising treating the solid support to recover the polynucleotide.

Claim 3

A method according to claim 2 wherein the solid support is treated with an elution solution to desorb the polynucleotide followed by separation of the elution solution from the solid support.

Claim 4

A method according to claim 3 wherein the elution solution comprises phosphate solution, pyrophosphate solution, tripolyphosphate solution, phytic acid solution or 50% formamide.

Claim 5

A method according to claim 3 or 4 wherein the polynucleotide is recovered from the elution solution.

Claim 9

A method according to claim 8 wherein the particles have a size of about 1 micron.

Claim 12

A method according to claim 8 or 9 wherein the particles are magnetic.

Claim 15

An assay for detecting a polynucleotide comprising a target sequence in a solution known or suspected to contain the polynucleotide which comprises: (a) bringing the solution into contact with an oligonucleotide probe for the target sequence which is smaller than the polynucleotide under hybridizing conditions in order to form a hybrid between the probe and target sequence, if present; (b) contacting the solution with a solid support having a plurality of cations available to interact with anions on the polynucleotide under conditions which binds said polynucleotide to said solid support but which are insufficient to bind said oligonucleotide to said solid support; wherein said solid support is capable of separating polynucleotides according to their charge or length and is: a magnetic amine solid support; a magnetic propylamine solid support; a magnetic quaternary ammonium solid support; a magnetic poly-D-lysine functionalized solid support; a poly-D-lysine functionalized polyurethane solid support; a spermine latex solid support; a Tris (2-amino ethyl) amine latex solid support; a Tris (2-amino ethyl) amine beaded agarose solid support; or a Tris (2-aminoethyl) polyurethane solid support. (c) detecting the amount of polynucleotide or probe either bound to the solid support or remaining in suspension as a qualitative or quantitative measure of said target sequence in the solution.

Claim 16

An assay for detecting a polynucleotide comprising a target sequence in a solution known or suspected to contain the polynucleotide which comprises: (a) contacting the solution with a solid support having a plurality of cations available to interact with anions on the polynucleotide under conditions which binds said polynucleotide to said solid support wherein said solid support is capable of separating polynculeotides according to their charge or length and is: a magnetic amine solid support; a magnetic propylamine solid support; a magnetic quaternary ammonium solid support; a magnetic poly-D-lysine functionalized solid support; a poly-D-lysine functionalized polyurethane solid support; a spermine latex solid support; a Tris (2-amino ethyl) amine latex solid support; a Tris (2-amino ethyl) amine beaded agarose solid support; or a Tris (2-aminoethyl) polyurethane solid support. (b) contacting the solid support and any polynucleotide bound thereto with a solution containing an oligonucleotide probe smaller than the polynucleotide under hybridizing conditions in order to form a hybrid between the probe and target sequence, if present, but which conditions are insufficient to bind said oligonucleotide probe to said solid support; and (c) detecting the amount of polynucleotide or probe either bound to the solid support or remaining in suspension as a qualitative or quantitative measure of said target sequence in the solution.

Claim 17

An assay according to claim 15 or 16 wherein the solid support is in the form of a suspension.

Claim 18

An assay according to any one of claims 15 to 17 wherein the oligonucleotide probe is labelled.

Claim 19

An assay according to any one of claims 15 to 18 wherein the detecting step (c) comprises: (i) separating the solid support from unhybridized probe; and (ii) detecting the presence of any probe thus separated and/or any probe bound to the polynucleotide on the solid support.

Claim 20

An assay according to any one of claims 15 to 18 wherein the detecting step (c) comprises: (i) separating the solid support from unhybridized probe; (ii) treating the solid support with an elution solution to remove any hybrid between the probe and target formed and/or any probe; and (iii) detecting the presence of probe thus eluted.

Claim 21

An assay according to claim 20 wherein the hybrid elution solution is 50% formamide, phosphate solution, pyrophosphate solution, tripolyphosphate solution or phytic acid solution.

Claim 22

An assay according to any one of claims 15 to 21 wherein the polynucleotide is at least 3 times larger than the oligonucleotide probe.

Claim 23

An assay according to claim 22 wherein the polynucleotides is at least 5 times larger than the oligonucleotide probe.

Claim 24

An assay according to any one of claims 15 to 23 wherein the probe is an analogue of DNA or RNA having a lower negative charge in the phosphate backbone than the phosphate diesters of native DNA or RNA.

Claim 25

An assay according to claim 24 wherein the analogue is a methyl phosphonate.

Claim 26

An assay according to any one of claims 15 to 25 wherein the solid support is washed prior to detection of the oligonucleotide probe.

Claim 27

An assay according to any one of claims 15 to 26 wherein the solid support comprises particles having a size of about one micron.

Claim 28

An assay according to claim 27 wherein the particles are magnetic.

Claim 29

An assay according to claim 27 or 28 wherein the particles are metal oxide, glass, polyamide, polyester, polyolefin, polysaccharide, polyglycol or polyaminoacid particles.

Claim 30

An assay according to any one of claims 15 to 26 wherein the solid support comprises fibers.

Claim 31

An assay according to any one of claims 15 to 26 wherein the solid support is a membrane.

Claim 32

An assay method according to any one of claims 15 to 31 wherein the cations are alkyl or aryl amines, or imines.

Claim 33

A kit for assaying for a polynucleotide comprising a target sequence comprising: (a) an oligonucleotide probe smaller than the polynucleotide for the target sequence; and (b) a solid support having a plurality of cations available to interact with anions on the polynucleotide under conditions which binds said polynucleotide to said solid support but which are insufficient to bind said oligonucleotide to said solid support; wherein said solid support is capable of separating polynucleotides according to their charge or length and is: a magnetic amine solid support; a magnetic propylamine solid support; a magnetic quaternary ammonium solid support; a magnetic poly-D-lysine functionalized solid support; a poly-D-lysine functionalized polyurethane solid support; a spermine latex solid support; a Tris (2-amino ethyl) amine latex solid support; a Tris (2-amino ethyl) amine beaded agarose solid support; or a Tris (2-aminoethyl) polyurethane solid support.

Claim 34

A kit according to claim 33 which further comprises an elution solution able to elute hybrid the polynucleotide from the solid support.

Claim 35

A kit according to claim 34 wherein the hybrid elution solution is 50% formamide, phosphate solution, pyrophosphate solution, tripolyphosphate solution or phytic acid solution.

Claim 36

A kit according to any one of claims 33 to 35 wherein the probe is selected so that the target polynucleotide is at least 3 times larger than the probe.

Claim 37

A kit according to claim 36 wherein the target polynucleotide is at least 5 times larger than the probe.

Claim 38

A kit according to any one of claims 33 to 37 wherein the probe is an analogue of DNA or RNA having a lower negative charge in the phosphate backbone than the phosphate diesters of native DNA or RNA.

Claim 39

A kit according to claim 38 wherein the analogue is a methyl phosphonate analogue.

Claim 40

A kit according to any one of claims 34 to 39 wherein the probe has a label to permit its detection.

Claim 41

A kit according to any one of claims 33 to 40 wherein the solid support comprises particles.

Claim 42

A kit according to claim 41 wherein the particles are about one micron in size.

Claim 43

A kit according to claim 41 or 42 wherein the particles are magnetic.

Claim 44

A kit according to any one of claims 41 to 43 wherein the particles are metal oxide, glass, polyamide, polyester, polyolefin, polysaccharide, polyglycol or polyaminoacid particles.

Claim 45

A kit according to any one of claims 33 to 40 wherein the solid support is comprised of fibers.

Claim 46

A kit according to any one of claims 33 to 40 wherein the solid support is a membrane.

Claim 47

A kit according to any one of claims 33 to 46 wherein the cations are alkyl or aryl amines or imines.