Methods and Systems for Molecular Fingerprinting

Abstract

This invention relates in general to a method for molecular fingerprinting. The method can be used for forensic identification (e.g. DNA fingerprinting, especially by VNTR), bacterial typing, and human/animal pathogen diagnosis. More particularly, molecules such as polynucleotides (e.g. DNA) can be assessed or sorted by size in a microfabricated device that analyzes the polynucleotides according to restriction fragment length polymorphism. In a microfabricated device according to the invention, DNA fragments or other molecules can be rapidly and accurately typed using relatively small samples, by measuring for example the signal of an optically-detectable (e.g., fluorescent) reporter associated with the polynucleotide fragments.

1 . A molecular fingerprinting method comprising the steps of: (a) identifying a target polynucleotide, (b) selecting at least one fragment of the target polynucleotide, wherein the fragment is a fixed distance from a restriction site, to generate a set of one or more polynucleotide fragments, and (c) designating some or all of the set of fragments as molecular fingerprint corresponding to the target polynucleotide. 
   
   
       2 - 3 . (canceled) 
   
   
       4 . A method for identifying a polynucleotide sample comprising the steps of: (a) identifying a target polynucleotide; (b) selecting at least one fragment of the target polynucleotide, wherein the fragment is a fixed distance from a restriction site, to generate a set of one or more polynucleotide fragments; (c) designating some or all of the set of fragments as a fingerprint corresponding to the target polynucleotide; (d) synthesizing one or more oligonucleotide probes to complement the set of polynucleotide fragments; (e) combining the probes, a polynucleotide sample, nucleotide triphosphates, and polymerase to synthesize at least one polynucleotide strand; (f) cutting the strands with restriction enzymes to yield a set of sample fragments of fixed length; and (g) comparing the set of sample fragments to the fingerprint. 
   
   
       5 - 15 . (canceled) 
   
   
       16 . A method for detecting a particular nucleic acid in a sample, which particular nucleic acid has at least one restriction site, and which method comprises: (a) contacting the sample with a primer that hybridizes to the particular nucleic acid a predetermined distance from the restriction site, a polymerase and a plurality of nucleotides, so that a complementary nucleic acid is synthesized from the primer at least to the restriction site; (b) contacting the complementary nucleic acid with a restriction enzyme under conditions capable of cutting the complementary nucleic acid at the restriction site; and (c) detecting a nucleic acid fragment having a particular length equal to the fixed distance, wherein the presence of the nucleic acid fragment in the sample indicates that the particular nucleic acid is present in the sample. 
   
   
       17 - 35 . (canceled) 
   
   
       36 . A method for detecting a particular nucleic acid in a sample, which particular nucleic acid has at least one restriction site, and which method comprises: (a) contacting the sample with a restriction enzyme under conditions capable of cutting the particular nucleic acid at the restriction site; (b) contacting the sample with a primer that hybridizes to the nucleic acid a predetermined distance from the cut at the restriction site, a polymerase and a plurality of nucleotides, so that a complementary nucleic acid fragment is synthesized; and (c) detecting the complementary to nucleic acid fragment, wherein the presence of the complementary nucleic acid fragment in the sample indicates that the particular nucleic acid is present in the sample. 
   
   
       37 - 55 . (canceled)