公开(公告)号：US06270963B1

公开(公告)日：2001-08-07

申请号：US08750232

申请日：1996-11-29

申请人： John K. Stevens ,  James M. Dunn ,  Denis Capatos ,  David E. Matthews

发明人： John K. Stevens ,  James M. Dunn ,  Denis Capatos ,  David E. Matthews

IPC分类号： C12Q168

CPC分类号： C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/6886 ,  C12Q2600/156 ,  C12Q2600/16

摘要： A hierarchy of at least two assay techniques is utilized in testing for disease-associated mutations. The first assay in the hierarchy is selected to provide a highly specific test for the existence of the disease-associated mutation, although the accuracy of the test need not be high. The final assay in the hierarchy is selected to provide a highly accurate and highly specific test for the existence of the disease associated mutation. Intermediate tests of progressively greater accuracy may also be included in the hierarchy. Once the hierarchy has been selected for a given mutation-associated disease, a patient sample is analyzed the patient sample using the first, lowest accuracy assay in the hierarchy. If the result of the first assay is negative for the presence of a disease-associated mutation, then the next assay in the hierarchy is performed. This process is repeated until the final assay has been performed on all samples which gave negative results when tested by all less-accurate assays in the hierarchy. The test may be used for diagnosis and targeted screening for p53 mutations and mutations in the RB1 gene.

摘要翻译： 使用至少两种测定技术的层次来测试疾病相关突变。 选择层次结构中的第一个测定法为疾病相关突变的存在提供高度特异性的测试，尽管测试的准确性不需要很高。 选择层次结构中的最终测定法为疾病相关突变的存在提供高度准确和高度特异性的检验。 渐进更高精度的中间测试也可能包含在层次结构中。 一旦为给定的突变相关疾病选择了层次结构，则使用层次结构中的第一种最低精度的分析方法对患者样本进行分析。 如果第一次测定的结果对于存在疾病相关突变是阴性的，则进行层次结构中的下一个测定。 重复此过程，直到对所有样品进行最终测定，当通过层次结构中的所有较不准确的测定进行测试时，会产生阴性结果。 该检测可用于诊断和靶向筛选p53突变和RB1基因突变。

公开(公告)号：US6013436A

公开(公告)日：2000-01-11

申请号：US699628

申请日：1996-08-19

申请人： May Hui ,  James M. Dunn ,  John K. Stevens ,  Denis Capatos ,  David E. Matthews

发明人： May Hui ,  James M. Dunn ,  John K. Stevens ,  Denis Capatos ,  David E. Matthews

IPC分类号： C07K14/47 ,  C07K16/18 ,  C12Q1/68 ,  C07H21/04 ,  C12P19/34

CPC分类号： C12Q1/6827 ,  C07K14/4703 ,  C07K16/18 ,  C12Q1/6858 ,  C12Q1/6886 ,  C12Q2600/16

摘要： The instant invention relates to methods and products for the diagnosis of mutations in the von Hippel-Lindau tumor suppressor gene. More specifically, the products are DNA oligonucleotides for use in amplifying and sequencing genomic DNA and kits including these oligonucleotides. The method of the invention relates to a hierarchical system for cost-effective diagnosis of von Hippel-Lindau tumor suppressor disease-associated mutations.

摘要翻译： 本发明涉及用于诊断von Hippel-Lindau肿瘤抑制基因突变的方法和产品。 更具体地，产物是用于扩增和测序基因组DNA的DNA寡核苷酸，包括这些寡核苷酸的试剂盒。 本发明的方法涉及用于von Hippel-Lindau肿瘤抑制因子相关突变的成本效益诊断的分层系统。

公开(公告)号：US6063567A

公开(公告)日：2000-05-16

申请号：US779916

申请日：1997-01-07

申请人： Brenda L. Gallie ,  James M. Dunn ,  John K. Stevens ,  May Hui

发明人： Brenda L. Gallie ,  James M. Dunn ,  John K. Stevens ,  May Hui

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

CPC分类号： C12Q1/6886 ,  C12Q2600/16

摘要： Reliable and cost effective testing for mutations in the RB1 gene can be accomplished by (a) quantitatively amplifying exons of the sample RB1 gene using primers complementary to intron regions flanking each exon; and (b) determining the lengths and/or quantities of the amplification products for each exon and comparing that length or quantity to the length or quantity of amplification products obtained when a wild-type RB1 gene is amplified using the same primers. Differences in length between an amplified sample exon and the corresponding amplified wild-type exon reflect the occurrence on an insertion or deletion mutation in the sample RB1 gene. Differences in quantity reflect the complete absence of an exon, or heterozygosity for a mutant exon. Next, the nucleic acid sequence of each exon found to contain an insertion or deletion mutation is determined, or of all exons in the event no insertion or deletion mutations are identified. Preferably, the amplification of the exons is multiplexed so that more than one exon is amplified in a single vessel using sets of primers which provide gene fragments of distinctive lengths when used to amplify a normal RB1 gene.

摘要翻译： 可以通过（a）使用与每个外显子侧翼的内含子区域互补的引物定量扩增样品RB1基因的外显子来实现对RB1基因突变的可靠和成本有效的测试; 和（b）确定每个外显子的扩增产物的长度和/或数量，并将该长度或数量与使用相同引物扩增野生型RB1基因时获得的扩增产物的长度或数量进行比较。 扩增样本外显子与相应的扩增野生型外显子之间的长度差异反映样品RB1基因插入或缺失突变的发生。 量的差异反映了突变外显子完全不存在外显子或杂合性。 接下来，确定发现含有插入或缺失突变的每个外显子的核酸序列，或者在没有鉴定插入或缺失突变的情况下所有外显子的核酸序列。 优选地，外显子的扩增被多重化，使得在用于扩增正常RB1基因时提供具有特异长度的基因片段的引物组在单个容器中扩增多于一个外显子。

公开(公告)号：US6071726A

公开(公告)日：2000-06-06

申请号：US765626

申请日：1996-12-27

申请人： Eleftherios Diamandis ,  James M. Dunn ,  John K. Stevens

发明人： Eleftherios Diamandis ,  James M. Dunn ,  John K. Stevens

IPC分类号： C12Q1/68 ,  C12P19/34 ,  C07H21/04 ,  G01N33/53

CPC分类号： C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/6886 ,  C12Q1/686 ,  C12Q1/6869 ,  C12Q2600/16

摘要： Rapid and cost effective diagnosis of p53 mutations of a sample of patients is achieved by employing a selected plurality of diagnostic tools, in a hierarchy of increasing accuracy and cost per tool, in which each tool detects essentially no false positives. Diagnostic tests that may be included among the plurality of tests selected include, in order of increasing accuracy and cost:(a) immunoassays,(b) analysis of DNA from a patient sample by quantitative amplification of p53 exons using amplification primers complementary to intron regions flanking each exon and examination of the length or quantity of each amplified fragment for nucleotide insertions or deletions relative to the normal p53 gene. Preferably, the amplification primers are multiplexed so that more than one DNA fragment is amplified in a single vessel, using sets of primers which provide gene fragments of distinctive lengths when used to amplify a normal p53 gene; and(c) analysis of DNA from a patient sample by DNA sequencing of the p53 gene beginning with the sequencing of those regions most likely to harbor point mutations, and proceeding to sequence regions less likely to harbor point mutations.

摘要翻译： PCT No.PCT / US95 / 08605 Sec。 371日期1996年12月27日第 102（e）日期1996年12月27日PCT提交1995年7月7日PCT公布。 公开号WO96 / 01909 日期1996年1月25日通过使用所选择的多种诊断工具，以提高每个工具的精度和成本的层次结构实现对患者样本的p53突变的快速和成本有效的诊断，其中每个工具基本上不检测到假阳性。 可以包括在所选择的多个测试中的诊断测试包括：提高准确性和成本的顺序：（a）免疫测定，（b）使用与内含子区域互补的扩增引物定量扩增p53外显子来分析来自患者样品的DNA 侧翼于每个外显子，并检查每个扩增片段的长度或数量，用于相对于正常p53基因的核苷酸插入或缺失。 优选地，扩增引物被多重化，使得在单个容器中扩增多于一个DNA片段，使用提供用于扩增正常p53基因的特异长度的基因片段的引物组; 和（c）通过对最可能存在点突变的那些区域的测序开始的p53基因的DNA测序从患者样品中分析DNA，并且进行到不太可能存在点突变的序列区域。

公开(公告)号：US5776767A

公开(公告)日：1998-07-07

申请号：US570994

申请日：1995-12-12

申请人： John K. Stevens ,  James M. Dunn ,  Gregory Dee ,  James W. Cassidy

发明人： John K. Stevens ,  James M. Dunn ,  Gregory Dee ,  James W. Cassidy

IPC分类号： G01N27/447 ,  G06F19/00 ,  C12M3/00

CPC分类号： G01N27/44782

摘要： A virtual DNA sequencer combines a plurality of individual DNA sequencers. Samples of DNA or other nucleic acid from subjects are prepared and allocated in real time to particular lanes or sets of lanes in electrophoresis plates of the individual sequencers, with records kept of the allocations. The data resulting from the electrophoresis runs is collected and collated according to the identities of the subjects. The individual sequencers are networked, and each individual sequencer is preferably equipped with a data buffer large enough to accommodate all or substantially all of a data run, thus protecting the virtual sequencer from loss of valuable data in the event that the network is disrupted for some portion of the time of the data run. In this way, a plurality of sequencers is virtually the same as a single sequencer with a very large number of tracks each of which can run for a much longer sequencing run than an individual sequencer.

摘要翻译： 虚拟DNA测序仪组合了多个单独的DNA测序仪。 来自受试者的DNA或其他核酸的样品被制备并实时分配到各个定序器的电泳板中的特定泳道或泳道组，并保存有分配记录。 根据受试者的身份收集和整理电泳运行产生的数据。 各个顺控程序被联网，并且每个单独的定序器优选地配备有足够大的数据缓冲器，以容纳所有或基本上所有的数据运行，从而在网络被中断的情况下保护虚拟定序器免于丢失有价值的数据 部分时间的数据运行。 以这种方式，多个定序器与具有非常大数量的轨道的单个定序器几乎相同，每个轨道可以运行比单个定序器更长的排序运行。

公开(公告)号：US5834189A

公开(公告)日：1998-11-10

申请号：US577858

申请日：1995-12-22

申请人： John K. Stevens ,  James M. Dunn ,  James Leushner ,  Ronald J. Green

发明人： John K. Stevens ,  James M. Dunn ,  James Leushner ,  Ronald J. Green

IPC分类号： C12N15/09 ,  C12Q1/68 ,  G01N27/447 ,  C12P19/34

CPC分类号： C12Q1/6858 ,  C12Q1/6869 ,  C12Q1/6881 ,  C12Q1/6883 ,  C12Q1/689 ,  G01N27/44721 ,  C12Q2600/156 ,  C12Q2600/16

摘要： The allelic type of a polymorphic genetic locus in a sample is identified by first combining the sample with a sequencing reaction mixture containing a polymerase, nucleotide feedstocks, one type of chain terminating nucleotide and a sequencing primer to form a plurality of oligonucleotide fragments of differing lengths, and then evaluating the length of the oligonucleotide fragments. As in a standard sequencing procedure, the lengths of the fragments indicate the positions of the type of base corresponding to the chain terminating nucleotide in the extended primer. Instead of performing and evaluating four concurrent reactions, one for each type of chain terminating nucleotide, however, the sample is concurrently combined with at most three, and preferably only one, sequencing reaction mixtures containing different types of chain terminating nucleotides. The information obtained from this test is evaluated prior to performing any additional tests on the sample. In many cases, evaluation of the positions of only a single base using one sequencing reaction will allow for allelic typing of the sample.

摘要翻译： 通过首先将样品与含有聚合酶，核苷酸原料，一种类型的终止核苷酸和测序引物的测序反应混合物合并，形成不同长度的多个寡核苷酸片段，来鉴定样品中多态性遗传位点的等位基因型 ，然后评估寡核苷酸片段的长度。 如在标准测序程序中，片段的长度表示对应于延伸引物中的链终止核苷酸的碱基类型的位置。 然而，代替执行和评估四种并发反应，每种类型的链终止核苷酸一个样品同时与至多三个，优选仅一个含有不同类型的链终止核苷酸的测序反应混合物组合。 在对样品进行任何额外的测试之前，先评估从该测试获得的信息。 在许多情况下，使用一个测序反应评估仅一个碱基的位置将允许样品的等位基因分型。

公开(公告)号：US5552283A

公开(公告)日：1996-09-03

申请号：US388381

申请日：1995-02-14

申请人： Eleftherios Diamandis ,  James M. Dunn ,  John K. Stevens

发明人： Eleftherios Diamandis ,  James M. Dunn ,  John K. Stevens

IPC分类号： G01N33/50 ,  C12N15/09 ,  C12Q1/68 ,  G01N33/53 ,  G01N33/566 ,  C07H21/02 ,  C07H21/04 ,  C12P19/34

CPC分类号： C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/6886 ,  C12Q2600/156 ,  C12Q2600/16

摘要： Rapid and cost effective diagnosis of p53 mutations of a sample of patients is achieved by employing a selected plurality of diagnostic tools, in a hierarchy of increasing accuracy and cost per tool, in which each tool detects essentially no false positives. Diagnostic tests that may be included among the plurality of tests selected include, in order of increasing accuracy and cost:(a) immunoassays,(b) analysis of DNA from a patient sample by quantitative amplification of p53 exons using amplification primers complementary to intron regions flanking each exon and examination of the length or quantity of each amplified fragment for nucleotide insertions or deletions relative to the normal p53 gene. Preferably, the amplification primers are multiplexed so that more than one DNA fragment is amplified in a single vessel, using sets of primers which provide gene fragments of distinctive lengths when used to amplify a normal p53 gene; and(c) analysis of DNA from a patient sample by DNA sequencing of the p53 gene beginning with the sequencing of those regions most likely to harbor point mutations, and proceeding to sequence regions less likely to harbor point mutations.

摘要翻译： 通过采用所选择的多种诊断工具，以提高每个工具的精度和成本的等级来实现对患者样品的p53突变的快速和成本有效的诊断，其中每个工具基本上不检测到假阳性。 可以包括在所选择的多个测试中的诊断测试包括：提高准确性和成本的顺序：（a）免疫测定，（b）使用与内含子区域互补的扩增引物定量扩增p53外显子来分析来自患者样品的DNA 侧翼于每个外显子，并检查每个扩增片段的长度或数量，用于相对于正常p53基因的核苷酸插入或缺失。 优选地，扩增引物被多重化，使得在单个容器中扩增多于一个DNA片段，使用提供用于扩增正常p53基因的特异长度的基因片段的引物组; 和（c）通过对最可能存在点突变的那些区域的测序开始的p53基因的DNA测序从患者样品中分析DNA，并且进行到不太可能存在点突变的序列区域。

公开(公告)号：US5550020A

公开(公告)日：1996-08-27

申请号：US271942

申请日：1994-07-08

申请人： Brenda L. Gallie ,  James M. Dunn ,  John K. Stevens

发明人： Brenda L. Gallie ,  James M. Dunn ,  John K. Stevens

IPC分类号： C12Q1/68 ,  C07H21/02 ,  C07H21/04 ,  C12P19/34

CPC分类号： C12Q1/6886 ,  C12Q2600/16

摘要： Reliable and cost effective testing for mutations in the RB1 gene can be accomplished by quantitatively amplifying exons of the sample RB1 gene using primers complementary to intron regions flanking each exon; and then determining the lengths and/or quantities of the amplification products for each exon and comparing that length or quantity to the length or quantity of amplification products obtained when a wild-type RB1 gene is amplified using the same primers. Differences in length between an amplified sample exon and the corresponding amplified wild-type exon reflect the occurrence of an insertion or deletion mutation in the sample RB1 gene. Differences in quantity reflect the complete absence of an exon, or heterozygosity for a mutant exon. Next, the nucleic acid sequence of each exon found to contain an insertion or deletion mutation is determined, or of all exons in the event no insertion or deletion mutations are identified. Preferably, the amplification of the exons is multiplexed so that more than one exon is amplified in a single vessel using sets of primers which provide gene fragments of distinctive lengths when used to amplify a normal RB1 gene.

摘要翻译： 可以通过使用与每个外显子侧翼的内含子区域互补的引物定量扩增样本RB1基因的外显子来实现RB1基因突变的可靠和成本有效的测试。 然后确定每个外显子的扩增产物的长度和/或数量，并将该长度或数量与使用相同引物扩增野生型RB1基因时获得的扩增产物的长度或数量进行比较。 扩增样本外显子与相应的扩增野生型外显子之间的长度差异反映了样本RB1基因中插入或缺失突变的发生。 量的差异反映了突变外显子完全不存在外显子或杂合性。 接下来，确定发现含有插入或缺失突变的每个外显子的核酸序列，或者在没有鉴定插入或缺失突变的情况下所有外显子的核酸序列。 优选地，外显子的扩增被多重化，使得在用于扩增正常RB1基因时提供具有特异长度的基因片段的引物组在单个容器中扩增多于一个外显子。

公开(公告)号：US5545527A

公开(公告)日：1996-08-13

申请号：US271946

申请日：1994-07-08

申请人： John K. Stevens ,  James M. Dunn

发明人： John K. Stevens ,  James M. Dunn

IPC分类号： G01N33/50 ,  C12N15/09 ,  C12Q1/68 ,  G01N33/53 ,  G01N33/566 ,  C07H21/04 ,  C12P19/34 ,  C12Q1/70

CPC分类号： C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/6886 ,  C12Q2600/156 ,  C12Q2600/16

摘要： A hierarchy of at least two assay techniques is utilized in testing for disease-associated mutations. The first assay in the hierarchy is selected to provide a highly specific test for the existence of the disease-associated mutation, although the accuracy of the test need not be high. The final assay in the hierarchy is selected to provide a highly accurate and highly specific test for the existence of the disease associated mutation. Intermediate tests of progressively greater accuracy may also be included in the hierarchy. Once the hierarchy has been selected for a given mutation-associated disease, a patient sample is analyzed the patient sample using the first, lowest accuracy assay in the hierarchy. If the result of the first assay is negative for the presence of a disease-associated mutation, then the next assay in the hierarchy is performed. This process is repeated until the final assay has been performed on all samples which gave negative results when tested by all less-accurate assays in the hierarchy. The test may be used for diagnosis and targeted screening for p53 mutations and mutations in the RB1 gene.

摘要翻译： 使用至少两种测定技术的层次来测试疾病相关突变。 选择层次结构中的第一个测定法为疾病相关突变的存在提供高度特异性的测试，尽管测试的准确性不需要很高。 选择层次结构中的最终测定法为疾病相关突变的存在提供高度准确和高度特异性的检验。 渐进更高精度的中间测试也可能包含在层次结构中。 一旦为给定的突变相关疾病选择了层次结构，则使用层次结构中的第一种最低精度的分析方法对患者样本进行分析。 如果第一次测定的结果对于存在疾病相关突变是阴性的，则进行层次结构中的下一个测定。 重复此过程，直到对所有样品进行最终测定，当通过层次结构中的所有较不准确的测定进行测试时，会产生阴性结果。 该检测可用于诊断和靶向筛选p53突变和RB1基因突变。

公开(公告)号：US08026819B2

公开(公告)日：2011-09-27

申请号：US11639857

申请日：2006-12-15

申请人： Jason August ,  John K. Stevens ,  Paul Waterhouse

发明人： Jason August ,  John K. Stevens ,  Paul Waterhouse

IPC分类号： G08B21/22 ,  G08B21/18

CPC分类号： G06K19/07767 ,  G06K19/07749 ,  G08B13/2417 ,  G08B13/242 ,  G08B13/2431 ,  G08B13/2434 ,  G08B13/2445 ,  G08B13/2448 ,  G08B13/2462 ,  G08B13/2477 ,  G08B13/2482

摘要： Passive tags use two antennas with only limited mutual coupling, one of which receives a power/clock field and the other of which receives a data signal. An area-reading antenna, or two or more antennas, are deployed to generate the power/clock field, from a base station. The base station, or active tags, or both, generate the data signals from time to time. This topology together with the use of low frequencies permits area reads, and permits small and economical passive tags, and further permits localization of a particular passive tag as being nearby to a particular active tag.

摘要翻译： 无源标签使用两个只有有限互耦的天线，其中一个接收功率/时钟域，另一个接收数据信号。 部署区域读取天线或两个或更多个天线以从基站产生功率/时钟字段。 基站或有源标签或两者都不时产生数据信号。 这种拓扑结构与低频率的使用一起允许区域读取，并允许小型且经济的无源标签，并且还允许将特定无源标签定位在特定活动标签附近。