Identification of molecular sequence signatures and methods involving the same

Abstract

Novel means and methods for analyzing hybridization data derived from hybridization assays between a target nucleic acid and differently sequenced polynucleotide probes involve selecting probe sets that define reference sequences for sequence signatures and deriving useful data about the nature of the target nucleic acid molecule based on its hybridization to the probes. The methods are useful for determining whether the target contains a nucleic acid or polypeptide sequence signature, whether the target encodes a member of a gene family, or whether the target is derived from one of any number of genes.

1 . A method for determining whether a target molecule has a sequence from a gene family member comprising: 
 providing a polynucleotide array comprising, for each of at least two different gene family members, a set of polynucleotide probes that define a reference nucleotide sequence from the gene family member;    generating hybridization data by performing a hybridization reaction between the target nucleic acid molecule and the probes in the sets and detecting hybridization between the target nucleic acid molecule and each of the probes in the sets; and    processing the hybridization data to determine whether the target nucleic acid has the reference sequence from one of the gene family members.    
     
     
         2 - 17 . (canceled)  
     
     
         18 . A method of hybridizing a microarray of oligonucleotides bound to a polymer adsorbed on a surface of a siliceous substrate with a nucleic acid material comprising the step of incubating the nucleic acid material with the microarray of oligonucleotides on the adsorbed polymer surface in a hybridization solution at a hybridization temperature ranging from about 55.degree. C. to about 70.degree. C. so as to hybridize the nucleic acid material, wherein the hybridization solution comprises a buffer composition that comprises a pH within a range of pH 6.4 to 7.5, a non-chelating buffering agent that maintains the pH within the pH range, and a monovalent cation in a monovalent cation concentration ranging from about 0.01 M to about 2.0 M.  
     
     
         19 . The method of  claim 18 , wherein in the step of incubating, the non chelating buffering agent is selected from a group consisting of 2-[N-morpholino]ethanesulfonic acid (MES), 3-(N-Morpholine)propanesulfonic acid (MOPS), piperazine-N,N′-bis(2-ethansulfonic acid (PIPES), Tris(hydroxymethyl)aminomethane hydrochloride (TRIS-HCl), Hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES), and N -Tris(hydroxymethyl)methylglycine (TRICINE).  
     
     
         20 . The method of  claim 18 , wherein in the step of incubating, the monovalent cation is selected from a salt consisting of one or more of LiCl, NaCl and KCl and the monovalent cation concentration ranges from about 0.1 M to about 2.0 M.  
     
     
         21 . The method of  claim 18 , wherein the adsorbed polymer surface comprises a polycationic polymer.  
     
     
         22 . The method of  claim 21 , wherein the polycationic polymer is selected from a group consisting of one or more of polyethylenediamine, poly-acrylamide, poly-L-arginine, poly-L-histidine, and poly-L-lysine.  
     
     
         23 . The method of  claim 18 , wherein in the step of incubating, the buffer composition further comprises a chelating agent selected from a group consisting of one or more of ethylenediaminetetraacetic acid (EDTA),trans-1,2-diaminocyclohexanetetraacetic acid (CDTA) and diethylenetriaminopentaacetic acid (DTPA) that has a chelating agent concentration of less than about 100.mu.M.  
     
     
         24 . The method of  claim 18 , wherein in the step of incubating, the buffer composition further comprises an ionic surfactant selected from a group consisting of one or more of sodium dodecyl sulfate (SDS), lithium lauryl sulfate (LLS), N-lauryl sarcoside, acylated polypeptides, linear alkybenzene sulfonates, lignin sulfonates, paraffin sulfonates, sulfosuccinate esters, alkylnaphthalene sulfonates, isethionates, alkanolamine condensates, and N-alkylpyrrolidones, and wherein the step of incubating comprises using a hybridization chamber, and the ionic surfactant is provided in an amount sufficient to wet surfaces of the hybridization chamber and loosen bubbles impinged on the surfaces of the hybridization chamber.  
     
     
         25 . The method of  claim 24 , wherein the amount of ionic surfactant is a surfactant concentration ranging from about 0.01% to about 0.2% (w/v).  
     
     
         26 . The method of  claim 18 , wherein the buffer composition has a total cation concentration of about 0.02 M to about 2.0 M.  
     
     
         27 . The method of  claim 18 , wherein in the step of incubating, the non chelating buffering agent is 2-[N-morpholino]ethanesulfonic acid (MES), the monovalent cation is LiCl, the monovalent cation concentration is greater than or equal to 300 mM, the pH is within the range of pH 6.6 to 6.8.  
     
     
         28 . The method of  claim 27  wherein in the step of incubating, the buffer composition further comprises one or both of a chelating agent ethylenediaminetetraacetic acid EDTA having a chelating agent concentration of about 50.mu.M, and an ionic surfactant selected from sodium dodecyl sulfate (SDS), lithium lauryl sulfate (LLS) having a surfactant concentration that ranges from about 0.02% to about 0.1% (w/v), and the buffer composition has a total cation concentration of about 750 mM.  
     
     
         29 . The method of  claim 18 , before the step of incubating, further comprising the step of combining the nucleic acid material with the buffer composition.  
     
     
         30 . The method of  claim 18 , after the step of incubating, further comprising the step of interrogating the hybridized microarray at a first location, the first location being a physical location either where the incubation of the microarray is performed or another location separate from the microarray incubation location.  
     
     
         31 . The method of  claim 30 , further comprising the step of transmitting data representing a result of the interrogation.  
     
     
         32 . The method of  claim 31 , further comprising the step of receiving the transmitted data at a second location, the second location being a physical location that is different from one or both of the first location where the microarray interrogation is performed and the microarray incubation location.  
     
     
         33 . The method of  claim 32 , wherein the first location is remote from the second location, the remote first location being physically separated from the second location.  
     
     
         34 . A method of performing a high temperature hybridization assay comprising the step of: 
 incubating a nucleic acid material with a microarray of oligonucleotides in a hybridization solution at a hybridization temperature ranging from about 55.degree. C. to about 70.degree. C. so as to hybridize the nucleic acid material,    wherein the microarray comprises a siliceous substrate with an adsorbed polymer surface and oligonucleotides bound to the adsorbed polymer surface, and    wherein the hybridization solution comprises a pH within a range of pH 6.4 and 7.5 and a buffer composition, the buffer composition comprising a non-chelating buffering agent that maintains the pH within the range and a monovalent cation having a monovalent cation concentration ranging from 0.01 M and 2.0 M.    
     
     
         35 . A method of hybridizing a microarray of oligonucleotides with a nucleic acid material comprising the step of: 
 incubating the nucleic acid material with the microarray of oligonucleotides in a hybridization solution at a hybridization temperature ranging from about 55.degree. C. to about 70.degree. C. so as to hybridize the nucleic acid material, the oligonucleotides being bound to a polymer coating adsorbed on a surface of a siliceous substrate, the adsorbed polymer coating being non-covalently bound to the siliceous substrate surface,    wherein the hybridization solution comprises a buffer composition that comprises a pH within a range of pH 6.4 to 7.5, a non-chelating buffering agent that maintains the pH within the pH range, and a monovalent cation in a monovalent cation concentration ranging from about 0.01 M to about 2.0 M.    
     
     
         36 . The method of  claim 35 , wherein the non chelating buffering agent is 2-[N -morpholino]ethanesulfonic acid (MES), the monovalent cation being LiCl, the monovalent cation concentration being greater than or equal to about 300 mM, the pH being within the range of about pH 6.6 to about 6.8, and wherein the adsorbed polymer coating is a polycationic polymer.  
     
     
         37 . A method of reducing surface degradation to a microarray of oligonucleotides during a high temperature hybridization assay comprising: 
 incubating a nucleic acid material with the microarray of oligonucleotides in a hybridization solution at a hybridization temperature ranging from about 55.degree. C. to about 70.degree. C. so as to hybridize the nucleic acid material, the oligonucleotides being bound to a polycationic polymer that is adsorbed to a surface of a siliceous substrate, the adsorbed polycationic polymer being non-covalently bound to the siliceous substrate surface, wherein the hybridization solution comprises a buffer composition that comprises a pH within a range of pH 6.4 to 7.5, a non-chelating buffering agent that maintains the pH within the pH range, and a monovalent cation in a monovalent cation concentration ranging from about 0.01 M to about 2.0 M.