公开(公告)号：WO2005047474A3

公开(公告)日：2006-09-14

申请号：PCT/US2004037407

申请日：2004-11-10

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M ,  HOLMLIN R ERIK ,  ZHANG HONGHUA ,  SHI CHUNNIAN

Inventor： CROTHERS DONALD M ,  HOLMLIN R ERIK ,  ZHANG HONGHUA ,  SHI CHUNNIAN

IPC: C12Q1/68 ,  C12N20060101 ,  C12P19/34

CPC classification number: C12Q1/682 ,  C12Q1/6825 ,  C12Q2525/307 ,  C12Q2531/125 ,  C12Q2525/161 ,  C12Q2565/607 ,  C12Q2563/113 ,  C12Q2537/125

Abstract: The present disclosure relates to methods of detecting nucleic acid hybridization by monitoring an output signal. Some advantageous embodiments include techniques which can augment the signal. One such technique involves catalytic detection, such as with a peroxidase or other enzyme. Another technique involves "on-chip" amplification to enlarge the nucleic acid after hybridization has occurred.

Abstract translation: 本公开涉及通过监测输出信号来检测核酸杂交的方法。 一些有利的实施例包括可以增加信号的技术。 一种这样的技术涉及催化检测，例如用过氧化物酶或其它酶。 另一种技术涉及“片上”扩增，以在杂交发生后扩增核酸。

公开(公告)号：WO2005049848A3

公开(公告)日：2007-06-14

申请号：PCT/US2004037472

申请日：2004-11-10

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M

Inventor： CROTHERS DONALD M

IPC: C12Q1/68 ,  C07H21/04 ,  C12P19/34 ,  C12Q20060101

CPC classification number: C12Q1/6837 ,  C12Q2565/543 ,  C12Q2531/125 ,  C12Q2521/531

Abstract: Disclosed herein are methods of destabilizing double-stranded nucleic acid hybridization using an enzyme comprising DNA N-glycosylase activity. Also disclosed herein is the detection of a double-stranded target DNA wherein the hybridization of duplex strands has been at least partially disrupted thereby permitting invasion of a probe strand. Also disclosed herein are methods of using an enzyme comprising DNA N-glycosylase activity to generate single-stranded circular nucleic acids.

Abstract translation: 本文公开了使用包含DNA N-糖基化酶活性的酶使双链核酸杂交失稳的方法。 本文还公开了双链靶DNA的检测，其中双链体的杂交已被至少部分地破坏从而允许侵入探针链。 本文还公开了使用包含DNA N-糖基化酶活性的酶以产生单链环状核酸的方法。

公开(公告)号：WO2005021717A3

公开(公告)日：2005-10-13

申请号：PCT/US2004027412

申请日：2004-08-23

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M

Inventor： CROTHERS DONALD M

IPC: C12N20060101 ,  C12P19/34 ,  C12Q1/68

CPC classification number: C12Q1/6832 ,  C12Q1/6846 ,  C12Q2549/125 ,  C12Q2533/107 ,  C12Q2525/301

Abstract: The present application provides methods and compositions for use in detecting the presence of a target nucleic acid in a sample. In some embodiments, the methods employ sequestering agents which specifically interact with complementary nucleic acids which will be ligated together if the target nucleic acid is present in the sample. Detection of a ligation product comprising the complementary nucleic acids indicates that the target nucleic acid is present in the sample.

Abstract translation: 本申请提供了用于检测样品中靶核酸存在的方法和组合物。 在一些实施方案中，所述方法使用螯合剂，所述螯合剂与互补核酸特异性相互作用，如果靶核酸存在于样品中，互补核酸将连接在一起。 检测包含互补核酸的连接产物表明靶核酸存在于样品中。

公开(公告)号：WO2004099433A3

公开(公告)日：2005-07-21

申请号：PCT/US2004013514

申请日：2004-04-30

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M ,  HOLMLIN ERIK R ,  ZHANG HONGHUA ,  SHI CHUNNIAN

Inventor： CROTHERS DONALD M ,  HOLMLIN ERIK R ,  ZHANG HONGHUA ,  SHI CHUNNIAN

IPC: C12Q1/68

CPC classification number: C12Q1/6825 ,  C12Q1/682 ,  C12Q1/6827 ,  C12Q1/6837 ,  C12Q2565/607 ,  C12Q2563/137 ,  C12Q2533/107 ,  C12Q2531/125 ,  C12Q2521/301 ,  C12Q2525/179 ,  C12Q2565/501 ,  C12Q2531/137 ,  C12Q2527/107

Abstract: The present disclosure provides methods and compositions for conducting an assay to detect nucleic acid hybridization in the presence of oxygen. In particular, ruthenium complexes having a reduction potential that does not coincide with the reduction potential of molecular oxygen are disclosed and amperometric techniques for their use are described. In preferred embodiments, the ruthenium complex is ruthenium (III) pentaamine pyridine and the nucleic acid hybridization event that is detected is DNA hybridization. Further, techniques for enhancing detectable contrast between hybridized and unhybridized nucleic acids are disclosed. In particular, the use of elongated target strands as well as the use of uncharged probe strands are discussed.

Abstract translation: 本公开提供用于进行测定以在氧存在下检测核酸杂交的方法和组合物。 特别地，公开了具有与分子氧的还原电位不一致的还原电位的钌络合物，并且描述了它们的使用的电流分析技术。 在优选的实施方案中，钌络合物是钌（III）五胺吡啶，并且所检测的核酸杂交事件是DNA杂交。 此外，公开了用于增强杂交和未杂交核酸之间的可检测对比度的技术。 特别地，讨论了使用细长的靶股以及使用不带电的探针股线。

公开(公告)号：WO2004099755A3

公开(公告)日：2004-12-23

申请号：PCT/US2004013222

申请日：2004-04-30

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M ,  HOLMLIN R ERIK ,  SHI CHUNNIAN

Inventor： CROTHERS DONALD M ,  HOLMLIN R ERIK ,  SHI CHUNNIAN

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

CPC classification number: C12Q1/6827 ,  C12Q1/682 ,  C12Q1/6825 ,  C12Q1/6837 ,  C12Q2533/107 ,  C12Q2531/125 ,  C12Q2521/301 ,  C12Q2525/179 ,  C12Q2565/501 ,  C12Q2531/137 ,  C12Q2527/107 ,  C12Q2565/607

Abstract: The present invention relates to the detection of somatic cell mutations, particularly as part of a method to screen for cancer or precancer. The disclosure includes techniques for extracting and isolating oligonucleotides from a patient and conducting hybridization assays. Preferred embodiments include a combination of the following steps: extracting a biological sample from a patient, purifying a nucleic acid from a biological sample, amplifying a nucleic acid, isolating a nucleic acid in single stranded form, cyclizing a nucleic acid, elongating a nucleic acid, controlling hybridization stringency, amplifying a nucleic acid on a chip, and detecting hybridization.

Abstract translation: 本发明涉及检测体细胞突变，特别是作为筛选癌症或癌前期的方法的一部分。 本公开包括用于从患者中提取和分离寡核苷酸并进行杂交测定的技术。 优选的实施方案包括以下步骤的组合：从患者提取生物样品，纯化来自生物样品的核酸，扩增核酸，分离单链形式的核酸，环化核酸，延长核酸 ，控制杂交严格性，扩增芯片上的核酸，并检测杂交。

公开(公告)号：WO2005010199A3

公开(公告)日：2009-04-09

申请号：PCT/US2004022465

申请日：2004-07-14

Applicant: GENEOHM SCIENCES INC ,  CROTHERS DONALD M ,  EIS PEGGY S

Inventor： CROTHERS DONALD M ,  EIS PEGGY S

IPC: C12Q1/68 ,  C07H21/02 ,  C12Q20060101

CPC classification number: C12Q1/6827 ,  C12Q1/6825 ,  C12Q1/6858 ,  C12Q2565/519 ,  C12Q2561/109 ,  C12Q2525/301 ,  C12Q2531/125 ,  C12Q2525/143 ,  C12Q2565/514

Abstract: A universal tag assay is disclosed wherein at least one invasive cleavage reaction (ICR) is used to generate tagged molecules having identifier tags corresponding to target nucleotide sequences, and further wherein hybridization of any tagged molecule with a complementary detection probe on a universal detector indicates the presence of the corresponding target in the sample being assayed. Preferred embodiments include the use of ICR to generate molecules suitable for use in the universal tag assay to detect variant nucleotide sequences including single nucleotide polymorphisms (SNPs), allelic variants, and splice variants. Hybridization of tagged molecules to detection probes is preferably detected by electrochemical readout, in particular the use of ruthenium amperometry to detect hybridization of identifier tags to detection probes immobilized on a universal detector, preferably a universal chip having gold or carbon electrodes.

Abstract translation: 公开了一种通用标签测定法，其中使用至少一种侵入性切割反应（ICR）产生具有与靶核苷酸序列相对应的标识符标签的标记分子，并且进一步地，任何标记分子与通用检测器上的互补检测探针的杂交表明 测定样品中相应靶标的存在。 优选的实施方案包括使用ICR产生适用于通用标签测定的分子，以检测包括单核苷酸多态性（SNP），等位基因变体和剪接变体的变体核苷酸序列。 标记分子与检测探针的杂交优选通过电化学读数检测，特别是使用钌电流分析法来检测标识符标签与固定在通用检测器上的检测探针的杂交，优选具有金或碳电极的通用芯片。

公开(公告)号：WO03091406A2

公开(公告)日：2003-11-06

申请号：PCT/US0312824

申请日：2003-04-22

Applicant: GENEOHM SCIENCES ,  CROTHERS DONALD M ,  KOENIGSBERGER CAROL

Inventor： CROTHERS DONALD M ,  KOENIGSBERGER CAROL

IPC: C12N15/09 ,  C12Q1/68 ,  C12N

CPC classification number: C12Q1/6853 ,  C12Q2565/519 ,  C12Q2525/204 ,  C12Q2521/301 ,  C12Q2531/113 ,  C12Q2525/131

Abstract: The present disclosure relates to methods for generating single-stranded DNA molecules of defined sequence and length. Specifically, a region of template containing target sequence is amplified by PCR or RCA, exogenous sequence is introduced by primers or probes used in amplification, double-stranded amplification products are converted to single-stranded amplification products, and single-stranded amplification products are trimmed to produce short single-stranded DNA molecules of defined sequence and length.

Abstract translation: 本公开涉及产生具有确定的序列和长度的单链DNA分子的方法。 具体地，通过PCR或RCA扩增含有靶序列的模板区域，扩增引物或探针引入外源序列，将双链扩增产物转化为单链扩增产物，并修饰单链扩增产物 以产生具有确定的序列和长度的短单链DNA分子。

公开(公告)号：WO2004044549A3

公开(公告)日：2004-10-21

申请号：PCT/US0335378

申请日：2003-11-05

Applicant: GENEOHM SCIENCES ,  CROTHERS DONALD M ,  HOLMLIN R ERIK

Inventor： CROTHERS DONALD M ,  HOLMLIN R ERIK

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

CPC classification number: C12Q1/6837 ,  C12Q1/682 ,  C12Q1/6825 ,  C12Q1/6827 ,  C12Q2533/107 ,  C12Q2531/125 ,  C12Q2521/301 ,  C12Q2525/179 ,  C12Q2565/501 ,  C12Q2531/137 ,  C12Q2527/107 ,  C12Q2565/607

Abstract: The present invention disclosure provides methods and compositions for a universal tag assay wherein a universal detector having detection probes is incubated with tagged molecules having identifier tags corresponding to targets, and hybridization of an identifier tag to a complementary detection probe indicates the presence of the corresponding target in the material being assayed. In particular, the invention disclosure provides methods and compositions for detecting target nucleotide sequences in a sample by target-dependent manipulations that generate tagged molecules having identifier tags corresponding to target nucleotide sequence, where incubation of tagged molecules with a universal detector having detection probes permits hybridization of identifier tags to complementary detection probes, thereby indicating the presence of the target nucleotide sequence corresponding to each identifier tag. Preferred embodiments include use of the universal tag assay for detecting variant sequences including single nucleotide polymorphisms (SNPs), allelic variants, and splice variants. Preferred embodiments further include the use of ruthenium amperometry to detect hybridization of tagged DNA or RNA molecules to detection probes immobilized on a universal detector, preferably a universal chip having gold or carbon electrodes.

Abstract translation: 本发明公开内容提供了用于通用标签测定法的方法和组合物，其中具有检测探针的通用检测器与具有对应于目标的标识符标签的标记分子一起温育，并且标识符标签与互补检测探针的杂交表明存在相应的靶标 在被分析的材料中。 特别地，本发明公开内容提供了用于通过靶标依赖性操作检测样品中的靶核苷酸序列的方法和组合物，所述靶标依赖性操作产生具有对应于靶核苷酸序列的标识符标签的标签化分子，其中用具有检测探针的通用检测器 的标识符标签到互补的检测探针，从而指示对应于每个标识符标签的目标核苷酸序列的存在。 优选实施方案包括使用通用标签测定法来检测包括单核苷酸多态性（SNP），等位基因变体和剪接变体的变体序列。 优选的实施方式进一步包括使用钌电流分析来检测标记的DNA或RNA分子与固定在通用检测器，优选具有金或碳电极的通用芯片上的检测探针的杂交。

公开(公告)号：WO2004016755A3

公开(公告)日：2004-08-26

申请号：PCT/US0325544

申请日：2003-08-14

Applicant: GENEOHM SCIENCES ,  KAWASHIMA TADASHI RYAN ,  HOLMLIN ERIK ,  CROTHERS DONALD M

Inventor： KAWASHIMA TADASHI RYAN ,  HOLMLIN ERIK ,  CROTHERS DONALD M

IPC: C07H19/00 ,  C07H21/02 ,  C07H21/04 ,  C12N20060101 ,  C12P19/34 ,  C12Q1/68

CPC classification number: C12Q1/6862 ,  C12Q1/682 ,  C12Q1/6844 ,  C12Q2531/137 ,  C12Q2531/125 ,  C12Q2521/301 ,  C12Q2561/101 ,  C12Q2531/113

Abstract: The present disclosure relates to methods for generating single-stranded DNA molecules of defined sequence and length using ligase chain reaction (LCR). Specifically, a region of template containing target sequence is amplified by LCR, exogenous sequence is introduced by LCR primers or probes used in amplification, and LCR products may be used in further amplification steps involving rolling circle amplification (RCA) or polymerase chain reaction (PCR). LCR products may include sequence complementary to the backbone of a padlock probe, where the LCR product hybridizes to a padlock probe and after ligation of the padlock, serves as polymerazation primer. After amplification, single-stranded amplification products are trimmed to produce short single-stranded DNA molecules of defined sequence and length.

Abstract translation: 本公开涉及使用连接酶链反应（LCR）产生具有确定的序列和长度的单链DNA分子的方法。 具体地，通过LCR扩增含有靶序列的模板区域，通过LCR引物或用于扩增的探针引入外源序列，并且LCR产物可用于进一步扩增步骤（RCA）或聚合酶链反应（PCR ）。 LCR产物可以包括与挂锁探针的主链互补的序列，其中LCR产物与挂锁探针杂交，并且在挂锁连接之后，用作聚合引物。 扩增后，修饰单链扩增产物以产生具有规定的序列和长度的短单链DNA分子。