Method for producing double-stranded DNA whose terminal homopolymer part is eliminated and method for determining nucleotide sequence
First Claim
1. A method for producing double-stranded DNA comprising:
- providing single-stranded DNA comprising a homopolymer at one or both ends;
employing said single-stranded DNA to produce a double-stranded DNA comprising a homopolymer at one or both ends;
wherein the double-stranded DNA does not contain internal type IIS restriction sites or is chemically modified in order to protect the double-stranded DNA from internal type IIS restriction enzyme cleavage;
after producing said double-stranded DNA, treating said double-stranded DNA comprising a homopolymer at one or both ends with a type IIS restriction enzyme to partly or fully eliminate at least one of the homopolymers, wherein the restriction enzyme is capable of cleaving said double-stranded DNA at a cleavage site separate from the recognition site therefor.
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Accused Products
Abstract
Disclosed is a method for producing double-stranded DNA comprising treating double-stranded DNA having a homopolymer part or parts at one or both ends with a restriction enzyme to partly or fully eliminate at least one of the homopolymer part or parts. The restriction enzyme is capable of cleaving double-stranded DNA at a cleavage site separate from a recognition site therefor. Disclosed is a method for determining a nucleotide sequence of double-stranded DNA utilizing one or both strands of the double-stranded DNA as a template, wherein the double-stranded DNA used as the template is double-stranded DNA prepared by the above method of the present invention. The present invention provides a method for producing double-stranded DNA a part or all of which homopolymer part, which may inhibit nucleotide sequence determination, is eliminated, and a method for determining a nucleotide sequence utilizing as a template the double-stranded DNA prepared by the above method of the present invention, whose homopolymer part is partially or fully eliminated.
7 Citations
19 Claims
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1. A method for producing double-stranded DNA comprising:
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providing single-stranded DNA comprising a homopolymer at one or both ends;
employing said single-stranded DNA to produce a double-stranded DNA comprising a homopolymer at one or both ends;
wherein the double-stranded DNA does not contain internal type IIS restriction sites or is chemically modified in order to protect the double-stranded DNA from internal type IIS restriction enzyme cleavage;
after producing said double-stranded DNA, treating said double-stranded DNA comprising a homopolymer at one or both ends with a type IIS restriction enzyme to partly or fully eliminate at least one of the homopolymers, wherein the restriction enzyme is capable of cleaving said double-stranded DNA at a cleavage site separate from the recognition site therefor. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19)
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Specification