公开(公告)号：US07205129B1

公开(公告)日：2007-04-17

申请号：US09514113

申请日：2000-02-28

申请人： Frank B. Dean ,  A. Fawad Faruqi

发明人： Frank B. Dean ,  A. Fawad Faruqi

IPC分类号： C12P19/34 ,  C12Q1/68 ,  C07H21/04

CPC分类号： C12Q1/6844 ,  C12Q1/6853

摘要： Disclosed are compositions and methods useful for reducing the formation of artifacts during nucleic acid amplification reactions. The method uses special oligonucleotides, referred to herein as template-deficient oligonucleotides, that cannot serve as a template for nucleic acid synthesis over part of their length. This prevents the oligonucleotides from serving as effective templates in the formation of artifacts. The disclosed method involves using a template-deficient oligonucleotide as at least one of the oligonucleotides (preferably a primer) in a nucleic acid amplification reaction, where the template-deficient oligonucleotide comprises one or more template-deficient nucleotides, preferably at or near the 5′ end of the template-deficient oligonucleotide. The disclosed method is useful for reducing artifacts in any nucleic acid amplification reaction involving oligonucleotides. In a preferred form of the method the nucleic acid amplification reaction does not involve thermal cycling. The disclosed method is effective at reducing non-cycle oligonucleotide-based artifacts. Also disclosed are kits useful for reducing artifacts in nucleic acid amplification reactions. The disclosed kits include a template-deficient oligonucleotide, wherein the template-deficient oligonucleotide comprises one or more template-deficient nucleotides, and a nucleic acid polymerase.

摘要翻译： 公开了用于在核酸扩增反应期间减少伪影形成的组合物和方法。 该方法使用特定的寡核苷酸，本文称为模板缺陷型寡核苷酸，其不能作为核苷酸合成部分长度的模板。 这可以防止寡核苷酸作为伪像形成的有效模板。 所公开的方法包括在核酸扩增反应中使用模板缺陷型寡核苷酸作为寡核苷酸（优选引物）中的至少一种，其中模板缺陷型寡核苷酸包含一个或多个模板缺陷型核苷酸，优选为或接近5 “模板缺陷寡核苷酸的末端。 所公开的方法可用于减少涉及寡核苷酸的任何核酸扩增反应中的伪像。 在该方法的优选形式中，核酸扩增反应不涉及热循环。 所公开的方法在减少基于非循环寡核苷酸的伪像方面是有效的。 还公开了可用于减少核酸扩增反应中的伪像的试剂盒。 所公开的试剂盒包括模板缺陷型寡核苷酸，其中模板缺陷型寡核苷酸包含一个或多个模板缺陷核苷酸和核酸聚合酶。

公开(公告)号：US06977148B2

公开(公告)日：2005-12-20

申请号：US09977868

申请日：2001-10-15

申请人： Frank B. Dean ,  Roger S. Lasken ,  Linhua Fang ,  A. Fawad Faruqi ,  Osama A. Alsmadi ,  Mark D. Driscoll ,  Seiyu Hosono

发明人： Frank B. Dean ,  Roger S. Lasken ,  Linhua Fang ,  A. Fawad Faruqi ,  Osama A. Alsmadi ,  Mark D. Driscoll ,  Seiyu Hosono

IPC分类号： C12Q1/68 ,  C12P19/34

CPC分类号： C12Q1/6844 ,  C12Q2527/125

摘要： Disclosed are compositions and a method for amplification of nucleic acid sequences of interest. The disclosed method generally involves replication of a target sequence such that, during replication, the replicated strands are displaced from the target sequence by strand displacement replication of another replicated strand. In one form of the disclosed method, the target sample is not subjected to denaturing conditions. It was discovered that the target nucleic acids, genomic DNA, for example, need not be denatured for efficient multiple displacement amplification. The primers used can be hexamer primers. The primers can also each contain at least one modified nucleotide such that the primers are nuclease resistant. The primers can also each contain at least one modified nucleotide such that the melting temperature of the primer is altered relative to a primer of the same sequence without the modified nucleotide(s). The DNA polymerase can be φ29 DNA polymerase.

摘要翻译： 公开了用于扩增感兴趣的核酸序列的组合物和方法。 所公开的方法通常涉及靶序列的复制，使得在复制期间，复制的链通过另一复制链的链置换复制而从靶序列位移。 在所公开的方法的一种形式中，目标样品不经历变性条件。 已经发现，靶核酸，例如基因组DNA不需要被变性以用于有效的多位置扩增。 所使用的引物可以是六聚体引物。 引物还可以各自含有至少一个修饰的核苷酸，使得引物是耐核酸酶的。 引物还可以各自包含至少一个修饰的核苷酸，使得引物的融解温度相对于没有修饰的核苷酸的相同序列的引物而改变。 DNA聚合酶可以是phi29 DNA聚合酶。

公开(公告)号：US06617137B2

公开(公告)日：2003-09-09

申请号：US09982212

申请日：2001-10-18

申请人： Frank B. Dean ,  Roger S. Lasken

发明人： Frank B. Dean ,  Roger S. Lasken

IPC分类号： C12P1934

CPC分类号： C12Q1/6844 ,  C12Q1/6806 ,  C12Q2527/125 ,  C12Q2527/119 ,  C12Q2537/143 ,  C12Q2531/119 ,  C12Q2525/125

摘要： Disclosed are compositions and a method for amplification of nucleic acid sequences of interest. The disclosed method generally involves replication of a target sequence such that, during replication, the replicated strands are displaced from the target sequence by strand displacement replication of another replicated strand. In one form of the disclosed method, the target sample is not subjected to denaturing conditions. It was discovered that the target nucleic acids, genomic DNA, for example, need not be denatured for efficient multiple displacement amplification. The primers used can be hexamer primers. The primers can also each contain at least one modified nucleotide such that the primers are nuclease resistant. The primers can also each contain at least one modified nucleotide such that the melting temperature of the primer is altered relative to a primer of the same sequence without the modified nucleotide(s). The DNA polymerase can be &phgr;29 DNA polymerase.

摘要翻译： 公开了用于扩增感兴趣的核酸序列的组合物和方法。 所公开的方法通常涉及靶序列的复制，使得在复制期间，复制的链通过另一复制链的链置换复制而从靶序列位移。 在所公开的方法的一种形式中，目标样品不经历变性条件。 已经发现，靶核酸，例如基因组DNA不需要被变性以用于有效的多位置扩增。 所使用的引物可以是六聚体引物。 引物还可以各自含有至少一个修饰的核苷酸，使得引物是耐核酸酶的。 引物还可以各自包含至少一个修饰的核苷酸，使得引物的融解温度相对于没有修饰的核苷酸的相同序列的引物而改变。 DNA聚合酶可以是phi29 DNA聚合酶。

公开(公告)号：US07074600B2

公开(公告)日：2006-07-11

申请号：US10272465

申请日：2002-10-15

申请人： Frank B. Dean ,  Roger S. Lasken ,  Linhua Fang ,  A. Fawad Faruqi ,  Osama A. Alsmadi ,  Mark D. Driscoll ,  Seiyu Hosono ,  Michele Wisniewski ,  Wanmin Song

发明人： Frank B. Dean ,  Roger S. Lasken ,  Linhua Fang ,  A. Fawad Faruqi ,  Osama A. Alsmadi ,  Mark D. Driscoll ,  Seiyu Hosono ,  Michele Wisniewski ,  Wanmin Song

IPC分类号： C12P19/34

CPC分类号： C12Q1/6844 ,  C12Q1/6806 ,  C12Q2527/125 ,  C12Q2527/119 ,  C12Q2537/143 ,  C12Q2531/119 ,  C12Q2525/125

摘要： Disclosed are compositions and a method for amplification of nucleic acid sequences of interest. The disclosed method generally involves replication of a target sequence such that, during replication, the replicated strands are displaced from the target sequence by strand displacement replication of another replicated strand. In one form of the disclosed method, the target sample is not subjected to denaturing conditions. It was discovered that the target nucleic acids, genomic DNA, for example, need not be denatured for efficient multiple displacement amplification. The primers used can be hexamer primers. The primers can also each contain at least one modified nucleotide such that the primers are nuclease resistant. The primers can also each contain at least one modified nucleotide such that the melting temperature of the primer is altered relative to a primer of the same sequence without the modified nucleotide(s). The DNA polymerase can be φ29 DNA polymerase.

摘要翻译： 公开了用于扩增感兴趣的核酸序列的组合物和方法。 所公开的方法通常涉及靶序列的复制，使得在复制期间，复制的链通过另一复制链的链置换复制而从靶序列位移。 在所公开的方法的一种形式中，目标样品不经历变性条件。 已经发现，靶核酸，例如基因组DNA不需要被变性以用于有效的多位置扩增。 所使用的引物可以是六聚体引物。 引物还可以各自含有至少一个修饰的核苷酸，使得引物是耐核酸酶的。 引物还可以各自包含至少一个修饰的核苷酸，使得引物的融解温度相对于没有修饰的核苷酸的相同序列的引物而改变。 DNA聚合酶可以是phi29 DNA聚合酶。

公开(公告)号：US06323009B1

公开(公告)日：2001-11-27

申请号：US09605192

申请日：2000-06-28

申请人： Roger S. Lasken ,  Frank B. Dean ,  John Nelson

发明人： Roger S. Lasken ,  Frank B. Dean ,  John Nelson

IPC分类号： C12P1934

CPC分类号： C12Q1/6844 ,  C12Q2537/143 ,  C12Q2531/125

摘要： Processes for the amplification of target DNA sequences in the form of single or double stranded circular DNA molecules, especially those present in colony and plaque extracts, using multiple specific and/or random sequence oligonucleotide primers are disclosed along with methods for detecting such amplified target sequences. A kit containing components for use in the invention is also described.

摘要翻译： 公开了使用多个特异性和/或随机序列寡核苷酸引物扩增单链或双链环状DNA分子，特别是存在于菌落和斑块提取物中的靶DNA序列的方法以及用于检测这些扩增的靶序列 。 还描述了包含用于本发明的组分的试剂盒。